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South Eastern Area Laboratory Services
DEPARTMENT OF
CLINICAL CHEMISTRY & ENDOCRINOLOGY
SEALS Pathology Service
Randwick Campus
Dynamic Function Protocol
DETAILS OF TESTS AVAILABLE AND
SPECIMENS REQUIRED
Compiled by:
Phoebe Stanford
Registrar
Dept of Clinical Chemistry and Endocrinology
SEALS North
South Eastern Area Laboratory Services
SEALS PATHOLOGY SERVICE
RANDWICK CAMPUS
TABLE OF CONTENTS
PATIENT PREPARATION……………………………………………………….. 4
PROCEDURE FOR USING THE GLUCOMETER…………………………….. 6
PANCREAS
DIABETES
GLUCOSE TOLERANCE TEST …………………………………………………………………………..7
GESTATIONAL DIABETES SCREENING………………………………………………………………..8
GTT FOR INSULIN RESISTANCE (PCOS)…………………………………………………………….10
MEAL TOLERANCE……………………………………………………………………………………….10
EXTENDED GLUCOSE TOLERANCE TEST……………………………………………………11
ANTERIOR PITUITARY
ANTERIOR PITUITARY FUNCTION
ITT………………………………………………………………………………………………..12
CUSHING'S DISEASE
OVERNIGHT DEXAMETHASONE TEST……………………………………………………15
BILATERAL SIMULTANEOUS INFERIOR PETROSAL SINUS SAMPLING…………….16
ACROMEGALY
GROWTH HORMONE SUPPRESSION TEST (OGTT) FOR ACROMEGALY……………26
POSTERIOR PITUITARY
DIABETES INSIPIDUS
WATER DEPRIVATION TEST………………………………………………………………..27
ADRENAL
ADRENAL INSUFFICIENCY
SHORT SYNACTHEN TEST…………………………………………………………………..30
SYNACTHEN TEST - CONGENITAL ADRENAL HYERPLASIA………………………….32
HYPERALDOSTERONISM
SALINE SUPPRESSION TEST………………………………………………………………..33
LYING/STANDING AND FRUSEMIDE TEST………………………………………………..34
ADRENAL VENOUS SAMPLING……………………………………………………………..36
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PHAEOCHROMOCYTOMA
CLONIDINE SUPPRESSION TEST…………………………………………………………..41
BONE
BONE BIOPSY PROTOCOL…………………………………………………………………………..42
APPENDICES
APPENDIX 1………………………………………………………………………………………………44
APPENDIX 2……………………………………………………………………………………………..45
APPENDIX 3………………………………………………………………………………………………..46
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PATIENT PREPARATION
All patients must be booked in through the endocrine nurse on extension 24680, or Teresa on
extension 24602 and be given the appropriate information regarding their test procedure.
Requests for the following dynamic function tests must have the lab request form faxed to Teresa
at the Diabetes centre on 9382 4688. Patient details are essential, including patient’ phone number
for contacting purposes.
1.
2.
3.
4.



Short synacthen test
Insulin tolerance test
Water deprivation test
Saline infusion test
All medication on Resuscitation trolley is checked, oxygen and suction equipment is
tested.
Calibrate glucose meter if required
Collect all medication, glucose drink and insulin as required.
All patients present to ambulatory care. The registered nurse greets the patient, introduces herself
and explains the procedure.

Check specimen tubes are correct.

Identify patient and check spelling of their name.

Check time written on each specimen.

Bulk bill with Medicare for Procedure; Additional bloods required separate Medicare
form

For payment of test follow SEALS Finance payment protocol
The patient is assessed by the registered nurse before performing the procedure. The assessment
includes:
Taking a short medical history
 Check level of patient anxiety - reassure the patient and position comfortable for
procedure
 Patient’s medication – establish time and dose of last medications. Some tests require
morning medication to be omitted.
 Time of fasting - check that special diet requirements have been followed.
 Known allergies
 Weight & Height
 Blood pressure and pulse

Note down on commenced time of 0 minute; time & dosage of medication
 Time of adverse reaction & noted what action was taken eg. Glucose dose etc.
All information is documented on the patients’ continuation notes.
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Choose the appropriate equipment to suit the vein, eg 21 g winged infusion set for cubital fossa, 23
g for smaller veins. A cannula may be inserted if attending nurse / medical officer feels it is
appropriate.







Three way tap is attached to the end of butterfly/cannula
Draw up normal saline in 20 mL syringe for line flushing
Attach syringe to 3-way tap.
Holder is also attached to 3-way tap for blood collection.
At completion of each timed specimen.
Flush line with saline frequently and use flush tube to draw up saline to clear 3 way
tap of blood
Draw off 2 mLs of blood before each collection and discard, to prevent contamination
with saline.
Lunch to be ordered and given to patient following the Insulin Tolerance Test
Review Date: This section was reviewed by Phoebe Stanford (Chemistry registrar) 2014
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PROCEDURE FOR USING GLUCOMETER
Equipment
Supplies to wash and dry patient’s finger
Gloves for operator
Accu-Chek Performa™ Meter
Accu-Chek Performa™ Test Strips with code chip (check expiry date)
Lancing Device
Gauze swab or cotton wool
Method
Operator to use gloves
Wash and dry patient’s finger
Prepare lancet device
Insert a test strip into meter in direction of arrows
Make sure the code number on display matches the code number on the test strip container
When flashing blood drop symbol appears of display, perform finger prick
Gently squeeze finger to assist blood flow
Touch the drop of blood to the front edge of the yellow window of the test strip
When you see “6” flash, you have enough blood
Your result will appear on the screen in 5 seconds
Dispose of blood soiled articles in blood contaminated waste
Dispose of lancet device in sharps container
Recommendations
1. Washing patient’s finger ensures there are no contaminates that can interfere with result e.g.
fruit juice.
2. Drying patient’s finger ensures that there is no thinning of the blood that will result in a low
reading.
3. Using the sides of the finger tips ensures less pain for the patient.
4. Using the depth settings on the Lancing Device can help lessen pain and bruising at puncture
site.
5. Using a warm wash on hands can aid blood flow to the fingers.
6. If blood glucose result is questionable perform a quality control test and repeat blood glucose
test.
QUALITY CONTROL CHECKS
For detailed instruction on quality control checks and procedure for performing a blood glucose test
refer to POWH Clinical Business rules Manual December 2011, Section 15: Diabetes
Blood Glucose Monitoring and Quality Control – Accu-Chek Performa
(http://seslhnweb/powh/documents/cpm/Section15/accucheckperforma.pdf)
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GLUCOSE TOLERANCE TEST
Indication:
Screening for diabetes mellitus
Procedure:
Patient to attend after an overnight fast.
Collect blood at baseline (time 0), and then administer 75g glucose drink over 5-10minutes. Blood
is collected 60 and 120 minutes after the glucose load.
The patient is required to fast (no food or drink) and remain resting until the final sample is
collected.
Blood collected:
Time
0 min
120 min (2 hr)
Glucose (Sodium
fluoride) – 1mL
*
*
Interpretation:
WHO 2006 Criteria
Fasting 0 min glucose
(mmol/L)
Normal
< 6.1
Increased risk for diabetes 6.1-6.9
(IFG or IGT)
Diabetes Mellitus
 7.0
120 min glucose
(mmol/L)
<7.8
7.8-11
 11.1
Review Date: Reviewed by Phoebe Stanford (Chemistry registrar) June 2015
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PREGNANCY DIABETES SCREENING
2 hrs PREGNANCY DIABETES SCREEN (PDS - 2hrs 75g GTT)
Indication:
Screening for gestational diabetes/ diabetes in pregnancy
Procedure:
Patient to attend after overnight fast
Collect blood at baseline (time 0), then administer 75g glucose drink. Blood collected 60 and 120
minutes after the glucose load. The patient is required to fast (no food or drink) and remain resting
until the final sample is collected.
Blood collected:
Time
0 min
60min (1 hr)
120 min (2 hr)
Glucose (Sodium fluoride) – 1mL
*
*
*
Interpretation:
Gestational diabetes mellitus should be diagnosed at any time during pregnancy if any of the
following criteria are met:
(a) Fasting plasma glucose 5.1–6.9 mmol/l
(b) 1-h post 75 g oral glucose load >10.0 mmol/l*
(c) 2-h post 75 g oral glucose load 8.5–11.0 mmol/l
*there are no established criteria for the diagnosis of diabetes based on the 1-h post-load value
Diabetes mellitus in pregnancy should be diagnosed by the 2006 WHO criteria for diabetes if any
of the following criteria are met:
(a) Fasting plasma glucose > 7.0 mmol/l
(b) 2-h plasma glucose > 11.1 mmol/l following a 75 g oral glucose load
(c) a random plasma glucose > 11.1 mmol/l in the presence of diabetes symptoms.
Notes:
 The above criteria for gestational diabetes have been based on the Hyperglycaemia and
Adverse Pregnancy Outcome (HAPO) study – an international multicentre cohort study of
25,505 pregnant women tested with a 2-h 75g OGTT and then followed through pregnancy to
detect primary and secondary outcomes.
 The diagnostic values were defined on the basis of an odds ratio of 1.75 for adverse neonatal
outcomes (birth weight >90th percentile, cord C-peptide >90th percentile, and neonatal percent
body fat >90th percentile) compared with mean values, for fasting plasma glucose, 1-hour, and
2-hour OGTT plasma glucose values.
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 Although the diagnostic criteria are based on the risk of adverse pregnancy outcomes, there is a
continuous risk of adverse outcomes with increasing glycaemia, and therefore diagnostic
thresholds are somewhat arbitrary.

Appropriate follow up is recommended for all patients with a positive result, including referral to
a diabetes educator and dietitian, and review by either Endocrinologist or Obstetric physician.
References:



Report of a World Health Organization Consultation Diagnostic criteria and classification of
hyperglycaemia first detected in pregnancy: A World Health Organization Guideline. Diabetes Research
and Clinical Practice 2014; 103: 341-63.
ADIPS Consensus Guidelines for the Testing and Diagnosis of Gestational Diabetes Mellitus in Australia
(modified June 2014)
http://adips.org/downloads/2014ADIPSGDMGuidelinesVJune2014FINALforWEB.pdf
HAPO Collaborative Research Group. Hyperglycaemia and adverse pregnancy outcomes. New Eng J
Med 2008; 358:1991-2002
Review Date: Reviewed by Phoebe Stanford (Chemistry registrar) December 2014
ONE HOUR PREGNANCY DIABETES SCREEN (PDS - 1 hr 50g GTT)
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GTT FOR INSULIN RESISTANCE / POLYCYSTIC OVARY SYNDROME (PCOS)
(2 hrs GTT – 3 days High Carbohydrate Diet)
Overnight fasting, no fluid allowed & resting
Time
BSL (Sodium fluoride)
– 1mL
*
*
*
O min
60 min
120 min
Insulin (Gold gel)
(if requested) – 2mL
*
*
*
REFERENCE RANGE
4 - 10 fold increase in insulin levels max at 30 - 60 mins, falling to fasting level or below by 2 - 4
hours.
MEAL TOLERANCE TEST
This procedure is a physiological stimulus for insulin release and may be useful in the evaluation of
diabetes and hypoglycaemia.
Meal consists of:
2 slices of white bread toasted
5g margarine on each slice of bread
250mL unsweetened orange juice
Meal eaten at time 0 and consumed within 15 minutes.
Collect blood specimens as follows:
Time
Glucose
Insulin
C-Peptide
Trigs
Spare
(gold gel)
-30
*
*
*
-15
*
*
*
*
*
0
*
*
*
*
*
15
*
*
*
*
30
*
*
*
*
*
60
*
*
*
*
*
90
*
*
*
*
120
*
*
*
*
*
150
*
*
*
180
*
*
*
*
*
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EXTENDED GLUCOSE TOLERANCE TEST
Patient is given a diet sheet for a high carbohydrate diet, which is to be followed for 3 days prior to
the test. Patient fasts from 2100hr the night before the test.
An indwelling needle suitable for blood withdrawal over a 2 - 3 hour period is inserted into a
suitable vein. Patient is required to be on complete bed rest throughout the procedure.
Collect "0 mins" pre-test specimen for glucose and insulin if requested. Check blood glucose with
glucose meter. Consultant to be contacted if fasting blood glucose level is > 7.8mmol/l.
Patient is given glucose to drink (1 bottle containing 75g glucose drink in 300 mL.).
Collect further blood specimens for BSL and Insulin for 3 hours. Commence timing after glucose
has been drunk.
Cannula is kept clear between sampling by flushing with Normal Saline. Draw out 2 ml of blood
and discard prior to each collection.
Time
0 min
60 min (1 hr)
120 min (2 hr)
180 min (3 hr)
Glucose (Sodium
fluoride) – 1mL
*
*
*
*
Insulin (Gold gel) – 2mL
*
*
*
*
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INSULIN TOLERANCE TEST
Background:
This procedure causes hypoglycaemia which results in a stress response with stimulation of the
pituitary secretion of hGH, ACTH and Prolactin. This test is useful for investigating disorders of
hypothalamic-pituitary function. Adequate hypoglycaemia is required to achieve a response.
Indications:
Assessment of central adrenal insufficiency and growth hormone deficiency.
SIDE EFFECTS
The Insulin Tolerance Test is potentially hazardous. Signs and symptoms of hypoglycaemia occur
after about 30 minutes (see appendix 2) during which time careful supervision is required.
IV glucose may need to be given if profound hypoglycaemia occurs.
corticosteroids and IV fluids may be necessary to reverse shock.
Additionally, the
CONTRAINDICATIONS
Ischaemic Heart Disease, seizure disorder.
Preparation:
 Patient advised to:
o Fast overnight (no food/drink after dinner, water allowed)
o Omit glucocoticoids from the evening before (Last dose hydrocortisone midday the
day before)
o Bring lunch (i.e. sandwich) to be eaten after hypoglycaemia achieved
o Arrange to be collected, or accompanied by a family member or friend as they will
not be allowed to drive for 2 hours after the test
 Ensure Actrapid Insulin available
 Ensure the resuscitation trolley is within easy reach and stocked with 50% dextrose and
hydrocortisone
 Ensure glucometer available and perform QC prior to test
 Ensure specimen tubes ready:
o 7 grey top fluoride oxalate tubes
o 1 purple top EDTA tube
o 7 gold top serum tubes
o Blood gas syringe for urgent confirmation of hypoglycaemia
Test Protocol:
 Weight patient in light street clothing (for calculation of insulin dose)
 Insert IV cannula with patient resting in bed. This should be in a position suitable for
withdrawing blood and administration of IV dextrose should this be required.
 Check baseline blood glucose with glucose meter to be used for calculation of insulin dose.
 Patient to remain in bed for the remainder of test.
 Baseline blood collected after 20-30 minutes rest for Cortisol, glucose, ACTH and GH,
followed by administration of IV neutral insulin (Actrapid®) (time 0)
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










Dose of insulin to be determined by requesting consultant, or by Registrar (using the table
below as a guide)
Whole blood glucose levels to be measured by glucose meter every 5 min following IV
insulin. The lowest glucose level following IV insulin is usually at 20-30 minutes.
Venous blood samples taken every 30 min for 120 min for measurement of GH and cortisol
Adequate hypoglycaemia defined as blood glucose (BG) concentration < 2·2 mmol/l
occurring in parallel with the development of hypoglycaemic symptoms (see appendix 2)
must be achieved. This should be confirmed by a laboratory glucose measurement with an
urgent VBG prior to treatment of hypoglycaemia (a fluoride oxalate (grey top) should also
be sent for glucose). The VBG (with request form) should be taken directly to the clinical
chemistry lab and delivered to the scientist on the blood gas analyser. The glucose result
should be communicated by phone to the doctor in attendance with the patient to minimise
delay.
If adequate hypoglycaemia is not reached by 30 minutes and at least two consecutive
glucose readings show an upward trend a second (0·15 units/kg) or third (0·2 units/kg)
dose of IV insulin can be administered in consultation with the requesting consultant
Where there is additional insulin given sampling is prolonged such that there must be at
least 3 specimens following adequate hypoglycaemia (the last sample to be collected at
least 90 minutes after hypoglycaemia achieved).
If patient develops symptoms of hypoglycaemia despite a glucose meter glucose
>2.2mmol/L then a VBG sample should be collected and analysed urgently on the blood
gas analyser (as indicated above)
Once adequate hypoglycaemia is achieved the patient can be given a glucose drink and
eat their lunch. Venous sampling is to continue as per the protocol.
If the symptoms of hypoglycaemia are severe the hypoglycaemia can be terminated by
administration of IV dextrose (10-20ml aliquots as required). Where possible Blood should be
collected prior to administration of IV dextrose or oral carbohydrate. Sampling should be
continued as per the protocol. There is a standing order for the administration of 50%
dextrose if required for management of symptomatic hypoglycaemia (see appendix 1 and
3).
At all times a doctor or nurse must be in attendance. A doctor should be present to
administer the insulin but can leave once glucose levels start to rise following
hypoglycaemia.
If a patient has an adrenal crisis they should receive IV 0.9% saline and hydrocortisone
100mg
Insulin Dose:
Low dose
Standard dose
Insulin –resistant dose
High dose
Condition
Cortisol deficiency (ITT performed
for GH deficiency)
Morning cortisol >100nmol/L and
BMI <30
Obese and or metabolic syndrome
with fasting BSL >5.5mmol/L
Active
acromegaly,
Cushing’s
syndrome or type 2 diabetes
Insulin dose (units/kg)
0.1
0.15
0.2
0.3
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NOTE: THE TIME AND DOSAGE OF INSULIN MUST BE RECORDED IN THE MEDICAL RECORD
AND INCLUDED ON THE REQUEST FORM.
Blood to be collected as follows:
Tube
Glucose
Fluoride Oxalate
Cortisol
Serum
GH
Serum
ACTH
EDTA (on ice)
0
√
√
√
√
30
√
√
√
60
√
√
√
90
√
√
√
120
√
√
√
Interpretation
The test can only be interpreted if adequate hypoglycaemia (glucose <2.2mmol/L) was achieved.
Cortisol
There is a lack of consensus on the cortisol level which defines an adequate response. The
concentration of peak cortisol measured is dependent on the assay used. In addition the value
chosen to represent the cut off point is a factor of pre-test probability and desired sensitivity vs
specificity. The clinician should interpret the result in the clinical context when making a diagnosis
of adrenal insufficiency.
Ideally there would be assay specific cut off points with associated sensitivity and specificity
reported however, given a lack of such data, most centres use a cut off derived from the literature
using outdated assays, commonly either 500 or 550nmol/L.
Close correlation has been shown between the peak cortisol during an ITT with the 30 minute
cortisol post 250microg synacthen. Studies of A study by El-Farhan et al of Cortisol response to
synacthen in healthy individuals have determined a 2.5th percentile for 30minute Cortisol of between
524nmol/L in females (not on OCP) and 574nmol/L in males with the Roche cortisol assay.
In 2015 Roche introduced a new cortisol assay, Roche Cortisol II. This assay uses a new
monoclonal antibody which has less cross reactivity than the current assay. The Cortisol II assay
has also been standardised to the newest reference material IRMM/IFCC 451(ID-GC/MS), and so
results are more in line with measurements by the reference method. Results with the cortisol II
assay are approximately 15-20% lower than those with the old roche cortisol assay.
Based on the correlation with the old roche assay, a normal peak cortisol response during an ITT
with the Roche cortisol II assay should be approximately 440 nmol/L. Additionally the above study
by El-Farhan et al also measured the cortisol with GC-MS with a recommended cut off of 420nmol/l
by this method.
Growth Hormone
With polyclonal RIA, the cut-off values for stimulated GH levels for diagnosing adult growth
hormone deficiency were established at levels between 3 and 5 microg/liter. Whether lower cut-offs
should be used with the newer, more sensitive, two-site assays has not been definitively
determined.
In a multicenter study which used a sensitive, immunochemiluminescent two site assay, Biller et al
found a serum GH cut-point of 5.1 microg/liter to have a high sensitivity (96%) and specificity
(92%) for GH deficiency. While a cut point of 3.3microg/L achieved 95% specificity.
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References
1. Sarlos & Inder Selective use of the insulin tolerance test to diagnose hypopituitarism Internal
Medicine Journal 43 (2013)
2. GonzaÂlbez et al. Establishment of reference values for standard dose short synacthen test (250
mg), low dose short synacthen test (1 mg) and insulin tolerance test for assessment of the
hypothalamo-pituitary-adrenal axis in normal subjects. Clinical Endocrinology (2000) 53, 199-204
3. El-Farhan et al. Method-specific serum cortisol responses to the adrenocorticotrophin test:
comparison of gas chromatography mass spectrometry and five automated immunoassays. Clinical
Endocrinology (2013) 78, 673–680
4. Klose et al. Factors Influencing the Adrenocorticotropin Test: Role of Contemporary Cortisol Assays,
Body Composition, and Oral Contraceptive Agents The Journal of Clinical Endocrinology &
Metabolism 92(4):1326–1333
5. Molitch et al. Evaluation and Treatment of Adult Growth Hormone Deficiency: An Endocrine Society
Clinical Practice Guideline. J Clin Endocrinol Metab 96: 1587–1609, 2011
6. B Biller eta l. Sensitivity and Specificity of Six Tests for the Diagnosis of Adult GH Deficiency. The
Journal of Clinical Endocrinology & Metabolism 87(5):2067–2079
Review Date: Reviewed by Phoebe Stanford (Chemistry registrar) February 2015
Revised 17/10/2015
OVERNIGHT DEXAMETHASONE SUPPRESSION TEST
Indication:
Investigation of cushings syndrome
In normal subjects, the administration of a supraphysiological dose of glucocorticoid results in
suppression of ACTH and cortisol secretion. In endogenous Cushing’s syndrome, there is a failure
of this suppression when low doses of the synthetic glucocorticoid dexamethasone are given.
Preparation:
The patient requires a prescription for dexamethasone tablet. No fasting is required.
Test Protocol:
The patient takes 1mg oral dexamethasone between 2300-2400. Blood is collected the following
morning between 0800 - 0900 for cortisol.
SIDE EFFECTS:
Possible mood disturbances and insomnia.
INTERPRETATION:
Available assays for the measurement of cortisol are not standardised and results for a single
sample measured in various assays may be quite different.
In addition, antibody-based immunoassays can be affected by cross-reactivity with cortisol
metabolites and synthetic glucocorticoids. In particular the Roche assay has been shown to have significant positive
bias as compared to the reference method of GC-MS in ICU and renal failure patients.
Specific cut off points for the Roche assay are not available.
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Various cut offs have been reported, depending on sensitivity vs specificity. A widely used normal
response is a serum cortisol < 140 nmol/L, however as this diagnostic criterion may misclassify up
to 15% of patients as negative a cut off of <50nmol/L has been advocated to enhance sensitivity.
Post dexamethasone serum cortisol <50 nmol/liter is reported to achieve a sensitivity of >95%,
with specificity of 80%. On the other hand specificity is reported to be > 95% with a diagnostic
threshold of 140 nmol/liter.
Cautions:
Oestrogen increases the cortisol-binding globulin (CBG) concentration. Because serum assays
measure total cortisol, false-positive rates for the overnight DST are seen in 50% of women taking
the oral contraceptive pill. Wherever possible, oestrogen-containing drugs should be stopped 6
weeks before testing.
Variable absorption and metabolism of dexamethasone may influence the result. Drugs (e.g.
Phenytoin, Carbamazepine) and alcohol may reduce dexamethasone levels through induction of
hepatic enzymatic clearance of dexamethasone mediated through CYP 3A4.
References:
L Nieman et al. The Diagnosis of Cushing’s Syndrome: An Endocrine Society Clinical Practice Guideline. J
Clin Endocrinol Metab 93: 1526–1540, 2008
A Dodd et al. The effect of serum matrix and gender on cortisol measurement by commonly used
immunoassays. Ann Clin Biochem 2014 51: 379
Review Date: Reviewed by Phoebe Stanford (Chemistry registrar) February 2015
BILATERAL SIMULTANEOUS INFERIOR PETROSAL SAMPLING
Indication
To distinguish between pituitary-dependent corticotrophin (ACTH) hypersecretion and ectopic
ACTH syndrome
Preparation
1. Participants comprise radiologist, anaesthetist, endocrinologist (or advanced endocrine
trainee or fellow), 2 other endocrine staff (resident, registrar, or laboratory staff), and the
Endocrine Laboratory
2. Procedure is performed in the Radiology Department, usually takes 1-2 hours and
preferably should occur in the morning to ensure that samples arrive at the Lab during
working hours
3. The Endocrine Lab should be notified of the date and time of the procedure in advance to
permit appropriate allocation of staff on the day and advise regarding correct collection
tubes and optimal delivery of samples from Radiology to the Lab
4. Corticotropin releasing hormone (CRH) is requested through Pharmacy, by completing a
Special Access Scheme (SAS) form. This should be ordered at least 2 weeks prior to the
procedure to ensure availability on the day of the procedure (see appendix 4 for example)
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5. Any medication used to block cortisol production, e.g. ketoconazole or aminoglutethimide,
should be withheld 3 days before the procedure, and may be recommenced immediately
after the procedure
6. On the day before the procedure –
a. assess 24 hour urine free cortisol, concluding on the morning of procedure
b. check FBC, coagulation studies, renal function, and take a group and hold
c. inform radiologist if patient takes warfarin, heparin or is allergic to iodine contrast
7. On the morning of the procedure –
a. patient should fast from breakfast, if procedure is performed in the morning
b. if patient takes metformin, this should be stopped, and recommenced 2 days
following the procedure if renal function has not deteriorated
c. check consent
d. bring an ampoule of CRH, appropriate collection tubes, an ice transport container
(for ACTH samples), and pathology request forms
Equipment
2* 1L normal saline (heparinized)
Peripheral angiography kit
1* large sterile plastic
2* sheath of choice (e.g. 6Fr)
2* 4Fr glide catheter – vertebral shape
0.035 145cm Bentson wire
1* 10ml LS syringe
2* tegaderm
2* 5ml LL syringe (discard blood)
5ml LL syringes for samples – open as required
Note
If microcatheters are required –
Splash bowl
3* 1L normal saline (heparinized) + 2 pressure bags
Microcatheters
2* 2ml LL syringe, 2* 1ml LL syringe
1* small metal bowel
1* 5 inch plastic bowel
1* pkt embo labels
2* large drape (back table)
1* filter
Procedure
1. Establish venous access bilaterally, usually via the femoral veins
2. Advance both catheters into bilateral high internal jugular veins (HIJV)
3. After confirming correct position of catheters, sample slowly (over 1-2 minutes)
simultaneously from both HIJV and a peripheral vein
4. Advance both catheters into inferior petrosal sinuses (IPS)
5. Perform venogram to confirm correct position of catheters and assess venous drainage of
both IPS
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6. After satisfactory catheter placement is confirmed, sample slowly (over 1-2 minutes)
simultaneously from both IPS and a peripheral vein
7. Administer corticotropin releasing hormone (CRH, ovine or human) as IV bolus peripherally
(1mcg/kg to maximal dose of 100mcg)
8. Repeat sampling, simultaneously from both IPS and a peripheral vein, at 2-3, 4-5, 9-10 and
14-15 minutes
9. At the end of sampling, confirm fluoroscopically that both IPS catheters remain in correct
positions
Each sample should be at least 2 mL
All IPS samples are assayed for ACTH and prolactin
HIJV samples need only be assayed for ACTH
All peripheral samples are assayed for ACTH
Peripheral samples taken simultaneously with IPS samples are also assayed for prolactin and
cortisol
All ACTH samples must be chilled immediately to prevent proteolytic degradation and brought to
the Lab as soon as possible
Take care in correctly labeling test tubes, each with the site and time of collection and tests to be
performed
References
1. Oldfield EH, Doppman JL el. Petrosal sinus sampling with and without corticotrophin-releasing
hormone for the differential diagnosis of Cushing’s syndrome. NEJM 1991; 325: 897-905
2. Kaltsas GA, Giannulis MG, Newell-Price JDC et al. A critical analysis of the value of simultaneous
inferior petrosal sinus sampling in Cushing’s Disease and the occult ectopic adrenocorticotropin
syndrome. JCEM 1999. 84: 487-492
3. Swearingen B, Katznelson L, Miller K et al. Diagnostic errors after inferior petrosal sinus sampling.
JCEM 2004. 89: 3752-3763
4. Royal Perth Hospital protocol for petrosal and cavernous sinus sampling. April 2004
Drafted by Dr Jeremy Hoang, advanced endocrine registrar, Prince of Wales hospital in
15.01.2008
Reviewed by Drs Jason Wenderoth, Chris White, Warren Kidson, Anne Poynten, Xuguang
Han
RESULT
Name and age
MRN
Date
Radiologist(s)
Sites of catheterization
Sites of sampling
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
How was catheter placement confirmed?

Was venography performed to assess venous anatomy?
Cavernous sinus (R/L)
Inferior petrosal sinus (R/L)
Internal jugular vein (R/L)

What was the stimulus and dose used?
CRH alone
CRH+desmopressin
Desmopressin alone

What and when were the anaesthetic agents and doses used?

Were there problems with sample handling and analysis?

What adverse events were observed?

Other comments
ACTH
Periphery
R IPS
L IPS
R HIJV
L HIJV
HIJV
0
2-3 min
4-5 min
9-10 min
14-15 min
Prolactin
Periphery
R IPS
L IPS
0
2-3 min
4-5 min
9-10 min
14-15 min
Cortisol
Periphery
0
2-3 min
4-5 min
9-10 min
14-15 min
IPS = Inferior petrosal sinus
LIJV = Low internal jugular vein
HIJV = High internal jugular vein
CRH = Corticotrophin releasing hormone
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INFERIOR PETROSAL SINUS SAMPLING: PROCEDURAL AND DIAGNOSTIC
CONSIDERATIONS
Jeremy Hoang, Advanced registrar in Endocrinology, 2008
Prince of Wales hospital
In distinguishing pituitary-dependent ACTH hypersecretion from the ectopic ACTH syndrome, the
inferior petrosal sinus sampling (IPSS) is the most direct biochemical test. Whilst its place in the
diagnostic algorithm is not universally established, there is substantial evidence that it has the
highest discriminatory accuracy in the distinction of these two conditions. Some series reported
specificity at or approaching 100%, and sensitivity 95% or above before CRH stimulation and at or
close to 100% after CRH (1,2,14). However, with time and experience, subsequent series have
documented both false negative and false positive results and much less accuracy (3). A number
of factors related to aspects of the procedural protocol and interpretation of diagnostic thresholds
significantly influence the sensitivity, specificity and predictive values of the IPSS. This overview
addresses these diagnostic aspects of the IPSS and provides the rationale for the accompanying
protocol
Procedural considerations
1. Sites of catheterization
Most pituitary ACTH-secreting microadenomas lie on one side of the pituitary gland, drain
unilaterally and may generate a dominant or single ACTH central-peripheral gradient. Therefore
IPPSS should be performed bilaterally (2,12). A number of series has demonstrated the higher
sensitivity for diagnosing pituitary-based disease associated with bilateral IPS catheterization
studies compared with single IPS catheterization, with or without corticotropin stimulation. In one
study of 128 patients, including 107 with Cushing’s disease (CD) and 6 with ectopic ACTH
secretion (EAS), the sensitivity of a stimulated maximal IPS/ peripheral ACTH gradient > 2 was
97% in patients who had bilateral catheterization studies compared with 89.5% in those with single
catheterization (2). In the study by Oldfield et al (1), basal maximal
(ie taken between right or left IPS samples) IPS/ peripheral gradient ≥ 2 and stimulated maximal
gradient ≥ 3 had a sensitivity of 95% and 100% respectively. However, the sensitivity of a single
randomly selected basal and stimulated IPS/ peripheral gradient fell to 80% and 86% respectively.
Similarly, in another series of 53 patients including 44 patients with confirmed CD, the sensitivity of
an unstimulated maximal IPS/peripheral gradient ≥ 2 was 82% compared with the sensitivity of a
lower IPS/ peripheral gradient of only 29% (4). These findings indicate that results of unilateral
sampling are only diagnostically useful if the IPS/ peripheral gradient is above the diagnostic
threshold and cannot exclude Cushing’s disease if it falls below that threshold.
Rates of successful bilateral catheterization varied among published series (73% (5) vs 97.5% (1)).
Kaltsas et al reported routine use of IPSS in all patients with ACTH-dependent Cushing’s
syndrome (CS) from 1985 to 1997 (2). The improvement in their success rate of bilateral
catheterization from 65.2% to 87.5% over this time suggests experience is a major factor.
Internal jugular venous (IJV) sampling is less sensitive for diagnosing pituitary disease due to the
large dilutional factor produced by non-pituitary venous return to the IJV (3). Kaltsas et al showed
that using the same central-peripheral gradient criterion of ≥ 2, pre- or post-CRH stimulation,
sensitivity was only 58% (2). In another study of 65 patients with proven CD and 11 patients with
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confirmed EAS, all of whom underwent both IPSS and IJVS with CRH stimulation on separate
days, at 100% specificity, the IPSS had a sensitivity of 94%, increased to 98% if patients with
abnormal venous drainage were excluded, compared with IJVS sensitivity of 83% (6). The high
specificity of IJVS shown in this and other studies (7), as well as its relative simplicity suggest that
it may be a reasonable alternative where access to or experience with IPSS is limited. A negative
IJVS result does not exclude CD.
Teramoto et al suggested routine selective catheterization and sampling of the cavernous sinus in
preference to the IPS in distinguishing pituitary from ectopic disease (8). However, a number of
subsequent studies have shown that compared with the IPSS, the cavernous sinus sampling
(CVS) does not have superior accuracy for diagnosing pituitary disease (9,10), has inferior
lateralizing accuracy (10, 11, 12) and is associated with transient cranial nerve palsies (11, 12).
2. Anatomy of venous drainage and the role of venography
The venous anatomy allows drainage of each half of the anterior pituitary gland into the ipsilateral
cavernous sinus and IPS. In a large series of 501 patients with proven CD all of whom underwent
bilateral IPSS before and after CRH stimulation, all 4 patients (0.8%) who had false negative
results were demonstrated to have hypoplastic or atrophic IPS ipsilateral to the confirmed ACTHsecreting adenoma (14). In each case retrograde venogram filled the ipsilateral cavernous sinus,
but contralateral retrograde venogram failed to demonstrate drainage of the hypopplastic IPS into
the internal jugular vein. Review of a subgroup of 100 consecutive patients showed that 18% had
unilateral atrophic IPS and 7% had bilateral atrophic IPS. A similar prevalence of hypoplastic IPS
was reported by Lefournier et al (11). The authors proposed that whilst the hypoplastic or plexiform
IPS demonstrated appropriate positioning of the catheter, it resulted in an altered venous drainage
pattern, possibly due to the obstruction by the ipsilateral catheter and may explain a negative
result. In particular, when the contralateral retrograde venogram demonstrates lack of filling of the
atrophic IPS and collateral flow to other venous systems, a false-negative result is possible. They
recommended that the finding of anomalous IPS venogram should lead to selective catheterization
and sampling of the cavernous sinus or alternative drainage routes demonstrated by contralateral
retrograde venogram.
Rarely, the IPS does not connect with the ipsilateral internal jugular vein. When this anatomy is
demonstrated, sampling is not possible and alternate drainage routes should be sought eg
vertebral venous system (1,15)
3. Timing of sampling
Kaltsas et al (2) showed that following CRH stimulation, IPS/ peripheral gradient > 2 was 97%
sensitive and 100% specific for CD. This was achieved at 5 minutes in all cases of CD. Maximal
IPS/ peripheral ratio was reached at 5 minutes in 90% of cases of CD, at 10 minutes in 9% and 15
minutes in 1%. Sampling earlier than 5 minutes was not performed. In another study of 281
patients in which IPSS was performed at baseline, and post CRH 2-3, 5 and 10, and in some
patients 15 to 20, and 30 minutes, Oldfield et al demonstrated that at diagnostic cut-offs of basal
IPS/ peripheral gradient ≥ 2 and stimulated gradient
≥ 3, diagnostic accuracy was highest for sampling at 2-3 minutes (100%), was less at baseline and
5 minutes post-CRH (96% and 97% respectively) and progressively declined with time from CRH
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administration (92% at 10 minutes, 88% at 15-20 minutes, and 86% at 30 minutes). In this study,
maximal IPS/ peripheral gradient occurred 2-3 minutes or 5 minutes after CRH administration in
89% of cases of CD. In one patient with surgically proven CD who underwent IPSS at our centre, a
stimulated
IPS/ peripheral gradient ≥ 3 was reached only at 10 minutes.
4. Complications
Oldfield et al performed 281 IPSS of which 278 was successful (274 involving bilateral
simulataneous IPSS) and reported groin haematomas in several patients as the only adverse
events (1). Kaltsas et al performed 128 IPSS and reported no major complication. 3 patients
developed groin haematomas. Transient ear pain or discomfort was thought to arise from the
sensitive periosteum of the jugular fossa and suggests that the catheter may be too unnecessarily
high in the IPS (13). Facial flushing has also been reported during selective IPS catheterization
(16). The available literature suggests an overall low risk of serious complications. In experienced
centres, the incidence of major morbidity has been reported to be 0.2% (17). These include
transient cranial nerve palsy, pulmonary embolism and deep venous thrombosis, right atrial wall
perforation and transient cardiac arrhythmias (11, 17). Some centres use a single intravenous
bolus of anticoagulant at the beginning of the procedure to prevent IPS or cavernous sinus
thrombosis, but this practice is debatable.
Diagnostic considerations
1. Diagnostic thresholds
Many studies used the diagnostic criteria of basal maximal (right or left) IPS/ peripheral gradient ≥
2 and CRH-stimulated maximal IPS/ peripheral gradient ≥ 3 as being diagnostic of CD. In the study
by Oldfield et al (1), 281 patients with CS underwent IPSS, of which bilateral sampling was
successful in 278 (99%). 246 patients (87.5%) had surgically confirmed CD, 20 (7.1%) had EAS,
and 11 (3.9%) had primary adrenal disease. Basal maximal (right or left) IPS/ peripheral gradient ≥
2.0 had a sensitivity of 95%, specificity of 100% and diagnostic accuracy of 96% for CD. CRHstimulated maximal IPS/ peripheral gradient ≥ 3.0 had a sensitivity of 100%, specificity of 100%
and diagnostic accuracy of 100%. In an analysis of pooled data derived from this and 13 other
series from 1985-2005, totaling 800 patients with CD and 124 with presumed or proven EAS, the
same combined diagnostic cut-offs had a sensitivity of 95% and specificity of 93% (17). Kaltsas et
al showed that IPS/ peripheral gradient > 2 before or after CRH stimulation had a sensitivity of 97%
and specificity of 100% for CD (2)
As discussed previously, sensitivity for CD is expected to be substantially diminished in single
IPSS studies (see ‘sites of catheterization’).
The studies by Oldfield et al (1) and Kaltsas et al (2) also demonstrate a number of important
differences in basal and CRH-stimulated ACTH levels and IPS/ peripheral gradients in CD, EAS
and primary adrenal disease:
1. Mean basal IPS ACTH was significantly higher in CD than in EAS, although there was an
overlap between the 2 conditions (269 +/- 43, range 13-1340 ng/L vs 75 +/- 15, range 28129 ng/L,
p < 0.05)
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2. Response to CRH stimulation:
a. In patients with CD, both IPS and peripheral ACTH levels significantly increased
and stimulated maximal IPS/ peripheral gradient was significantly higher than the
basal maximal gradient (55.8 +/- 7.5 vs 9.5 +/- 1.4, p < 0.05)
b. In EAS, Oldfield et al showed a small but significant rise in maximal IPS ACTH
levels (55 +/- 19 pmol/L vs 49 +/- 18 pmol/L, p = 0.03) but no change in peripheral
ACTH levels, while Kaltsas showed no change in either IPS or peripheral ACTH
levels. Both showed no significant change in IPS/ peripheral gradient
c. In primary adrenal disease, Oldfield et al showed no significant change in maximal
IPS ACTH levels, and no change in maximal IPS/ peripheral gradient in those with
detectable peripheral ACTH levels
Swearingen et al (3) proposed that CRH-stimulated peripheral ACTH levels be used in addition to
the IPS/ peripheral gradient to improve diagnostic accuracy. In their study of 179 patients with CS,
of whom 139 had proven CD, 9/139 (6.5%) failed to meet diagnostic criteria (basal IPS/ peripheral
gradient ≥ 2, and stimulated gradient ≥ 3). However, 8 of these had > 35% increase in peripheral
ACTH levels after given CRH.
2. Diagnostic errors
In addition to anomalous IPS or cavernous sinus venous drainage (see ‘anatomy of venous
drainage and role of venography’), other explanations for a false negative result may include:
1. Basal sampling occurs in absence of a pulsatile spontaneous ACTH secretory episode
2. Insufficient ACTH response to CRH stimulation occurs, although this is probably rare
False positive results are rare but have been reported in the following situations:
1. Mild or cyclic CS, where the degree or absence of hypercortisolaemia fails to adequately
suppress the normal pituitary corticotroph population (11,18)
2. Ectopic CRH secretion causing pituitary corticotroph hyperplasia and high IPS ACTH levels
(11,19,20)
3. CRH and desmopressin
In a recent study by Tsagarakis et al of 54 patients with ACTH- dependent C (47 CD and 7
histologically confirmed EAS), the combination of CRH (100µg) and desmopressin (10µg) had a
sensitivity of 97.9% and specificity of 100% (21). However, while none of the patients with ectopic
disease met the diagnostic criteria (IPS/ peripheral gradient > 2), 5 of these 7 patients
demonstrated a significant rise in peripheral ACTH levels, likely due to stimulation of the ectopic
tumour by desmopressin. In the study by Findling et al, 2 patients with EAS, who underwent IPSS
with combination CRH and desmopressin stimulation, had IPS/ peripheral gradients > 2 (22). The
concern arises as to whether combination CRH and desmopressin stimulation is more likely than
CRH alone to cause the false positive results that occur with CRH stimulation. This remains to be
seen, pending further studies with greater numbers of patients with ectopic disease.
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4. Lateralization
The importance of the anatomy of venous drainage in interpreting lateralization IPSS result is
demonstrated in the study by Lefournier et al in which 86 patients underwent bilateral IPS and
cavernous sinus sampling, 69 of these underwent transphenoidal surgery and 58 cases of
microadenomas were identified (11). Basal peak intersinus gradient ≥ 1.4 was considered to
predict the side of the pituitary microadenoma, and a gradient < 1.4 was indicative of a midline
lesion. In the 49 patients in whom the location of a microadenoma predicted by the intersinus
ACTH gradient could be compared with histopathologic findings, the lateralizing accuracy was only
57% overall, but improved to 71% when both venograms and catheter placement were symmetric
(35 patients). In the latter subgroup, accuracy was 86% in patients with both catheters in the IPS
compared with 50% in those with both catheters in the cavernous sinuses. An intersinus gradient <
1.4 was only 13% accurate for a midline tumour.
The variable results of other series also underscored the unreliability of lateralizing studies using
IPSS
(65-86%, average 75%) and cavernous sinus sampling, with some demonstrating superiority to
MRI and others inferiority (11,23). Similarly the site of sampling (cavernous sinus vs IPS) and CRH
stimulation had inconsistent influence on the lateralizing accuracy (2, 11) and their importance is
debatable
REFERENCE
1.Edward H, Oldfield MD, Doppman JL el. Petrosal sinus sampling with and without corticotrophin-releasing
hormone for the differential diagnosis of Cushing’s syndrome. NEJM 1991; 325: 897-905
2. Kaltsas GA, Giannulis MG, Newell-Price JDC et al. A critical analysis of the value of simultaneous inferior
petrosal sinus sampling in Cushing’s Disease and the occult ectopic adrenocorticotropin syndrome. JCEM
1999; 84: 487-492
3. Swearingen B, Katznelson L, Miller K et al. Diagnostic errors after inferior petrosal sinus sampling. JCEM
2004; 89: 3752-3763
4. Wiggam MI, Heaney AP, McILRATH EM et al. Bilateral inferior petrosal sinus sampling in the differential
diagnosis of adrenocorticotropin-dependent Cushing’s syndrome: a comparison with other diagnostic tests.
JCEM 2000; 85: 1525-1532
5. Bonelli FS, Huston J, Carpenter PC et al. Adrenocorticotropic hormone-dependent Cushing’s syndrome:
sensitivity and specificity of inferior petrosal sinus sampling. Am J Neuroradiol 2000; 21: 690
6. Ilias I, Chang R, Pacak K et al. Jugular venous sampling: an alternative to petrosal sinus sampling for the
diagnostic evaluation of adrenocorticotropic hormone-dependent Cushing’s syndrome. JCEM 2004; 89: 3795
7. Erickson D, Huston J III, Young WF et al. Internal jugular vein sampling in adrenocorticotropic hormonedependent Cushing’s syndrome: a comparison with inferior petrosal sinus sampling. Clinical Endocrinology
2004; 60: 413-419
8. Teramoto A, Niemoto S, Takakura K et al. Selective venous sampling directly from cavernous sinus in
Cushing's syndrome. JCEM 1993;76:637-641
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9. Graham KE, Samuels MH, Nesbit GM et al. Cavernous sinus sampling is highly accurate in distinguishing
Cushing’s disease from the ectopic adrenocorticotropin syndrome and in predicting intrapituitary tumour
location. JCEM 1999; 84: 1602
10. Doppman JL, Nieman LK, Chang R et al. Selective venous sampling from the cavernous sinuses is not a
more reliable technique than sampling from the inferior petrosal sinuses in Cushing’s syndrome. JCEM 1995;
80: 2485-2489.
11. Lefournier V, Martinie M, Vasdev A et al. Accuracy of bilateral inferior petrosal or cavernous sinus
sampling in predicting the lateralization of Cushing’s disease pituitary microadenoma: influence of catheter
position and anatomy of venous drainage. JCEM 2003; 88: 196-203
12. Liu C, Lo J, Dowd C et al. Cavernous and inferior petrosal sinus sampling in the evaluation of ACTHdependent Cushing’s syndrome. Clinical Endocrinology 2004; 61: 478-486
13. Doppman JL, Oldfield E, Krudy AG et al. Petrosal sinus sampling for Cushing’s syndrome: anatomical
and technical considerations. Radiology 1984; 150: 99-103
14. Doppman JL, Chang R, Oldfield EH et al. The hypoplastic inferior petrosal sinus: a potential source of
false-negative results in petrosal sampling for Cushing’s disease. JCEM 1999; 84: 533-540
15. Miller DL, Doppman JL, Chang R. Anatomy of the junction of the inferior petrosal sinus and the internal
jugular vein. Am J Neuroradiol 1993;14:1075-1083
16. Lopez J, Barcelo B, Lucas T et al. Petrosal sinus sampling for diagnosis of Cushing’s disease: evidence
of false negative results. Clin Endocrin 1996; 45(2): 147-156
17. Uptodate 2007
18. Yamamoto Y, Davis DH, Nippoldt TB et al. False-positive inferior petrosal sinus sampling in the diagnosis
of Cushing's disease. Report of two cases. J Neurosurg 1995; 83:1087-1091
19. Young J, Denau C, Grino M et al. Pitfall of petrosal sinus sampling in a Cushing's syndrome secondary to
ectopic adrenocorticotropin-releasing hormone (ACTH-CRH) secretion. JCEM 1998; 83:305-308
20. Case Records of the Massachusetts General Hospital. Case 52. NEJM 1987; 317:1648-1658
21. Tsagarakis S, Vassiliadi D, Kaskarelis S et al. The application of the combined corticotrophin-releasing
hormone plus desmopressin stimulation during petrosal sinus sampling is both sensitive and specific in
differentiating patients with Cushing’s disease from the occult ectopic adrenocorticotropin syndrome. JCEM
2007; 92: 2080- 2086
22. Findling JW, Raff H. Newer diagnostic techniques and problems in Cushing’s disease. Endocrinol Metab
Clin North Am 1999; 28: 191-210
23. Newell-Price J, Trainer P, Besser M et al. The diagnosis and differential of Cushing’s syndrome and
pseudoCushing’s states. Endocr Rev 1998; 19: 647-672
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Growth Hormone suppression test (OGTT) for Acromegaly
Indication:
Diagnosis of acromegaly, and monitoring of response to treatment.
Preparation:
 Patient to fast overnight (no food/drink after dinner, water allowed)
 Ensure specimen tubes ready:
o 6 Gold top serum tubes
o 6 grey top fluoride oxalate tubes
Test Protocol:
 Take blood sample for GH, IGF-1 and glucose at time 0.
 Administer 75 grams oral glucose in water over 5-10 minutes.
 Take blood for GH and glucose at time 30, 60, 90 and 120 minutes.
Glucose
GH
IGF1
tube
Fluoride oxalate (grey top)
Serum (gold top)
Serum (gold top)
0
√
√
√
30
√
√
60
√
√
90
√
√
120
√
√
Interpretation:
In normal individuals, GH levels fall following oral glucose. Failure of suppression or a paradoxical
rise in GH suggests acromegaly. The cut off point depends on the assay used. With the Immulite
assay a nadir of >1 microg/L suggests active acromegaly. As there is good correlation between the
Roche and Immulite assays at this level, this cut off can also be used with Roche.
GH may fail to suppress in chronic renal failure, liver failure, active hepatitis, anorexia nervosa,
malnutrition, hyperthyroidism, diabetes and adolescence.
References:
P Freda. Monitoring of acromegaly: what should be performed when GH and IGF-1 levels are discrepant?
Clinical Endocrinology (2009) 71, 166–170
Arafat, A.M., Mohlig, M., Weickert, M.O. et al. (2008) Growth hormone response during oral glucose
tolerance test: the impact of assay method on the estimation of reference values in patients with acromegaly
and in healthy controls, and the role of gender, age, and body mass index. Journal of Clinical Endocrinology
and Metabolism, 93, 1254–1262.
Review Date: Reviewed by Phoebe Stanford (Chemistry registrar) February 2015
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WATER DEPRIVATION TEST
The aim of this investigation is to diagnose and, if necessary, separate the three entities central
diabetes insipidus, nephrogenic diabetes insipidus, and primary polydipsia.
PRE-TEST
The requesting doctor is required to inform Clinical Chemistry department on x29073 & CSR on
x29092 with patient’s detail and provide doctor’s contact number for staff to inform results.
1.
2.
3.
4.
Liaise with Clinical Chemistry staff for interpretation of the plasma osmolality test.
The patient is on their usual diet.
No fluids are allowed after 2400 h (midnight).
No breakfast is eaten on the morning of the test. The patient arrives at POWH at 0730
where a "dry" breakfast will be provided.
THE TEST
PHASE 1
_______________________________________________________________________
Time Weight
Urine
Blood
_______________________________________________________________________
volume
osmolality
osmolality
U/E/C
ADH
_______________________________________________________________________
0800 [0 h]
x
x
x
x
x
x
0900 [1 h]
x
x
x
x
1000 [2 h]
x
x
x
x
1100 [3 h]
x
x
x
x
1200 [4 h]
x
x
x
x
x
x
_______________________________________________________________________
When the results of all the urine and plasma osmolalities are available, the Endocrine Registrar
should be consulted as to whether the test needs to be continued, either as regards phase 2 or 3.
The Registrar may need to liaise with the relevant Staff Specialist/VMO concerning this.
Criteria for termination of test
If urine osmolality > 800 mmol/kg or urine osmolality / plasma osmolality > 3.0, diabetes insipidus
has been excluded and test is terminated.
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Criteria for proceeding directly to Phase 3 (see over).
1.
Weight loss >5% of initial weight.
OR
2.
Plasma osmolality >300 mmol/kg.
CONTINUATION OF TEST
PHASE 2
_______________________________________________________________________
Time
Weight
Urine
Blood
_______________________________________________________________________
volume osmolality
osmolality
U/E/C ADH
_______________________________________________________________________
1300 [5 h]
x
x
x
x
1400 [6 h]
x
x
x
x
1500 [7 h]
x
x
x
x
1600 [8 h]
x
x
x
x
x
x
_______________________________________________________________________
At conclusion of phase 2 proceed to phase 3 at this time unless urine osmolality > 800 mmol/kg or
urine osmolality / plasma osmolality > 3.0, in which case diabetes insipidus has been excluded and
test is terminated.
If plasma osmolality > 300 mmol/kg or weight loss > 5% of initial weight, proceed directly to phase
3 regardless of whether phase 2 has been completed.
PHASE 3
This part of the water deprivation test is carried out to exclude nephrogenic diabetes insipidus.
It is carried out, after consultation with the Endocrine Registrar, at any time during the day. For this
test give 5 units (0.25mL) aqueous vasopressin subcutaneously or, if this is not available, 40 μg
(0.04 mL) desmopressin intranasally. One hour later the patient should be weighed, and urine
collected for measurement of volume, osmolality, and sodium. Blood should also be collected at
that time for osmolality, electrolytes and antidiuretic hormone. The test is then terminated.
SUPERVISION
1.
2.
3.
Bed rest required.
No water should be available.
Patient should be supervised at all times, especially when urinating.
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WATER DEPRIVATION TEST (cont.)
INTERPRETATION OF TEST
The following is meant as a guide only.
_______________________________________________________________________
Urine osmolality (mmol/kg)
_______________________________________________________________________
After dehydration
After desmopressin
Diagnosis
_______________________________________________________________________
> 800
> 800
normal
< 300
> 800
central diabetes insipidus
< 300
< 300
nephrogenic diabetes insipidus
300 - 800
< 800
partial central diabetes insipidus,
nephrogenic diabetes insipidus or
primary polydipsia
______________________________________________________________________
Plasma sodium >145 AND plasma osmolality > 300 mmol/kg
Primary polydipsia excluded
Urinary osmolality/plasma osmolality < 1.5
Primary polydipsia excluded
Urinary osmolality/plasma osmolality > 3.0
Diabetes insipidus excluded
OR
urine osmolality > 800 mmol/kg
_______________________________________________________________________
NOTES
If the patient has a history of significant nocturia/polyuria, then water deprivation should be under
supervision in the hospital starting at 0800. It is the responsibility of the physician ordering the test
to advise that this procedure is required.
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SYNACTHEN TEST (ACTH STIMULATION TEST)
Indication:
This procedure tests adrenocortical reserve and is useful in the diagnosis of adrenal insufficiency.
Preparation:
Test to be performed in the morning.
Glucocorticoids (e.g Prednisone, Hydrocortisone, cortisone acetate) ideally to be withheld for 24
hours prior to the test. Last dose of Hydrocortisone may be given in the early afternoon prior to the
test.
No fasting is required.
Collect “0 mins” pre-test specimen for cortisol and ACTH. ACTH to be collected in EDTA tube on
ice and sent immediately to the laboratory.
RMO to give synacthen IMI.
adults:
children:
10 years
250microg
125microg
After injection of synacthen, collect further blood specimens:
Time (mins)
Cortisol (Gold gel)
Volume (mL)
2
0
*
ACTH (optional)
(EDTA)
2
*
30
*
60
*
SIDE EFFECTS
Nausea, palpitation or hot flushes which pass quickly. Local irritation and pain can occur at the
injection site.
Interpretation:
There is a lack of consensus on the cortisol level which defines an adequate response. The
concentration of peak cortisol measured is dependent on the assay used. In addition the value
chosen to represent the cut off point is a factor of pre test probability and desired sensitivity vs
specificity. The clinician should interpret the result in the clinical context when making a diagnosis
of adrenal insufficiency.
In a study performing ….non-OCP women respectively).
Ideally there would be assay specific cut off points with associated sensitivity and specificity
reported however, given a lack of such data, most centres use a cut off derived from the literature
using outdated assays, commonly either 500 or 550nmol/L. There is also a lack of consensus as to
whether the 30 minute of 60 minute value should be used.
A study by El-Farhan et al of Cortisol response to synacthen in healthy individuals determined a
2.5th percentile for 30 minute Cortisol of 524nmol/L in females (not on OCP) and 574nmol/L in
males with the Roche cortisol assay.
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In 2015 Roche introduced a new cortisol assay, Roche Cortisol II. This assay uses a new
monoclonal antibody which has less cross reactivity than the current assay. The Cortisol II assay
has also been standardised to the newest reference material IRMM/IFCC 451(ID-GC/MS), and so
results are more in line with measurements by the reference method. Results with the cortisol II
assay are approximately 15-20% lower than those with the old roche cortisol assay.
Based on the correlation with the old roche assay, a normal 30minute response with the Roche
cortisol II assay should be approximately 440 nmol/L. Additionally the above study by El-Farhan et
al also measured the cortisol with GC-MS with a recommended cut off of 420nmol/l by this method.
In another study ….Immulite assays respectively.
The above study did not
measure the 60 minute response, however other studies by Klose et al and GonzaAlbez et al have
demonstrated that this is higher than the 30minut time point. In the study by GonzaÂlbez et al,
using healthy volunteers with cortisol measured by a direct chemiluminiscence immunoassay the
5th percentile was 537nmol/L at 30minutes and 649nmol/L at 60minutes. This should be taken into
account when considering the 60minute cortisol level.
These studies did not perform a 60 minute cortisol measurement which has been shown to be higher.
1. El-Farhan et al. Method-specific serum cortisol responses to the adrenocorticotrophin test:
comparison of gas chromatography mass spectrometry and five automated immunoassays. Clinical
Endocrinology (2013) 78, 673–680
2. Klose et al. Factors Influencing the Adrenocorticotropin Test: Role of Contemporary Cortisol Assays,
Body Composition, and Oral Contraceptive Agents The Journal of Clinical Endocrinology &
Metabolism 92(4):1326–1333
3. GonzaÂlbez et al. Establishment of reference values for standard dose short synacthen test (250
mg), low dose short synacthen test (1 mg) and insulin tolerance test for assessment of the
hypothalamo-pituitary-adrenal axis in normal subjects. Clinical Endocrinology (2000) 53, 199-204
Review Date: Reviewed by Phoebe Stanford (Chemistry registrar) February 2015
Revised: 17/10/2015
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SYNACTHEN TEST FOR CONGENITAL ADRENAL HYPERPLASIA (CAH)
The synacthen test may be used in the diagnosis of CAH due to 21-hydroxylase deficiency.
In this case blood will also be collected for 17OHP at times 0, 30 and 60 minutes, and the test
should be performed in the follicular phase in female patients.
Time (mins)
Cortisol (Gold gel)
ACTH (optional)
(EDTA)
17OHP (Gold gel)
Volume (mL)
2
2
0
*
*
30
*
60
*
2
*
*
*
SHORT SYNACTHEN TEST – CONGENITAL ADRENAL HYPERPLASIA
SPECIAL PROTOCOL
For further characterisation of adrenal hyperplasia not due to 21-hydroxylase deficiency additional
steroid hormones may be measured.
Time
(minutes)
0
Gold
gel
* x3
30
* x3
60
* x3
90
* x3
Hormones
Cortisol; Testosterone; Androstenedione; DHEA; DHEAS;
17OHPregnenolone; 11-Deoxycortisol; Aldosterone; Progesterone
Cortisol; Testosterone; Androstenedione; DHEA; DHEAS;
17OHPregnenolone; 11-Deoxycortisol; Aldosterone; Progesterone
Cortisol; Testosterone; Androstenedione; DHEA; DHEAS;
17OHPregnenolone; 11-Deoxycortisol; Aldosterone; Progesterone
Cortisol; Testosterone; Androstenedione; DHEA; DHEAS;
17OHPregnenolone; 11-Deoxycortisol; Aldosterone; Progesterone
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SALINE INFUSION TEST (INTRAVENOUS SALINE LOAD TEST)
Indications for Test:
Same as for Oral Salt Load Test. This test is contraindicated in patients with congestive heart
failure or inability to tolerate high volume fluid infusions.
This test is only performed when indicated by raised ARR (Aldosterone Renin Ratio) and also
referred to the pre-test checklist listed below.
Procedure:







Consider obtaining informed consent.
Prior to the test check patient’s blood pressure.
Discontinue interfering drugs as in Screening Tests for Mineralcorticoid Excess.
After ensuring that the patient is off as many antihypertensives as possible for at least one
week and diuretics for 2 weeks. Infuse 2 litres of normal saline i.v. over 4 hours while the
patient is supine.
Check patient’s blood pressure during the test.
EUC at baseline
Obtain serum Aldosterone and Renin at baseline, 2 hours & 4 hours later.
Collect blood specimen as follow:
Time (hours)
0
2
4
Renin
(EDTA not on ice) 4 mL
*
*
*
Aldosterone
(Gold gel) 4mL
*
*
*
EUC
(Gold gel) 4mL
*
RELEVANT INFORMATION:
False negatives by stimulating rennin
 Dietary salt restriction
 Renovascular hepertension or malignant hepertension
 Pregnancy
 Treatment with diuretics (including spironolactone)
 Dihydropyridine calcium blockers
 Angiotension converting enzyme inhibitors
 Angiotension receptor antagonists
False negatives by suppressing aldosterone or affecting its clearance
 Selective serotonin reuptake inhibitors lower the ratio
 Uncorrected hypokalemia because potassium regulates aldosterone
False positives by suppressing rennin
 Beta-blockers
 Alpha-methyldopa
 Clonidine
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




Nonsteroidal anti-inflammatory drugs
False positives may occur in patients with renal dysfunction or advancing age
Females (have higher ratios than males)
Luteal menstrual phase (think fluid retention & bloating)
Oral ethylestradiol / drospirenone (but not implanted subdermal etonogestrel)
contraceptive, if calculated using direct rennin concentration (SEALS uses a direct rennin)
and not plasma rennin activity).
PRE-TEST CHECK LIST & INSTRUCTIONS
1. Pre-menopausal women to be checked in the follicular phase; exclude pregnancy.
2. Diuretics should be ceased at least 6 weeks and other interfering medications at least 2
weeks before ARR measurement substituting non-interfering agents (e.g.verapamil slowrelease +/- hydralazine and prazosin or doxazosin) where required.
3. Hypokalemia should be corrected and a liberal salt diet encouraged.
4. Collecting blood midmorning from seated patients following 2-4hrs upright posture improves
sensitivity.
5. The ARR is a screening test only and should be repeated once or more before deciding
whether to proceed to confirmatory suppression testing.
6. Liquid chromatography-tandem mass spectrometry aldosterone assays represent a major
advance towards addressing inaccuracies inherent in other available methods.
Factors affecting the aldosterone/rennin ratio:Stowasser, M. Ahmed, AH. Pimenta, E.Taylor, PJ. Gordon, RD
Hormone & Metabolic research. 44(3): 170-6, 2012 Mar.
LYING/STANDING AND IV FRUSEMIDE TEST (RENAL PROTOCOL)
Indications:
This test is performed for the investigation of primary hyperaldosteronism, when indicated by
raised ARR (Aldosterone Renin Ratio) AND the absence of interfering agents.
Patient preparation:
1. Stop all antihypertensive and diuretic medications for at least three weeks.
2. If severe hypertension is present then the patient may continue to achieve partial control
with alpha-blockers (eg.Prazosin) as these agents will interfere least with the test.
Protocol:
1. On arrival, the patient is instructed to lie flat in bed.
2. A butterfly needle or cannula is inserted in an appropriate vein – flush cannula with 5ml
N/Saline.
3. Once the patient has been lying flat for 60 minutes, draw blood for plasma aldosterone and
renin levels (discard first 5 mls of blood drawn). Flush cannula again with 5ml N/Saline.
4. Label sample and blood form Sample 1
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5. The patient is asked to rise and walk around for the next 30 minutes (the patient should not sit
in a chair).
6. After 30 mins of activity, again draw blood for plasma aldosterone and renin levels (discard first
5mls drawn), flush the cannula
7. Label samples and blood form Sample 2
8. Administer 20mg Frusemide intravenously (no faster that 4mg/min) and flush cannula.
9. After 30mins draw blood for plasma aldosterone and renin levels (discard first 5 mls drawn)
10. Label samples and blood form Sample 3
11. Carefully label all samples with MRN, name, date and time drawn and sequence number and
deliver to the Clinical Chemistry/ Endocrinology lab at SEALS, Level 4 Campus Centre POWH.
12. Attention Endocrinology lab should be written on each of the blood forms and the lab should be
contacted (24805) that the specimens are on their way
If above test result is positive then there is a need to exclude Glucocorticoid-suppressible
Hyperaldosteronism by:
1. Prescribing Dexamethasone 0.5 mg qid for four days
2. Then repeat Aldosterone and Renin levels
3. Label sample with MRN, name, date and “post dexamethasone” and deliver to Endocrinology
lab, SEALS, level 4, Campus Centre
Specimen collection:
Specimen
Time (minutes)
1
2
3
0
30
60
Aldosterone –
Renin - EDTA
serum (gold tube) (purple tube) NOT
on ice
*
*
*
*
*
*
Note: Renin sample should NOT be collected on ice
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Adrenal Venous Sampling
1. OBJECTIVE:
The most useful technique in the investigation of primary aldosteronism is adrenal venous
sampling. Blood samples are obtained from both adrenal veins to analyse aldosterone levels;
hormone level ratios are compared with each other and with the level in the inferior vena cava
below the adrenal veins.
The patient will be adequately prepared for the Adrenal Venous Sampling and receive optimal care
during and after the procedure.
2. PRINCIPLES OF ACTION:
Adrenal Venous Sampling will take place under the following principles: Sterile technique (ACORN
Standard) Infection control (Infection control policy) Informed consent (Consent policy) Correct
Patient, Correct Procedure, Correct Site Policy Radiation safety. The procedure is documented
appropriately
3. DEFINATIONS:
Digital Subtraction Angiography
Is a type of imaging technique utilising Fluoroscopy as a guide to obtain images, clearly visualising
blood vessels in a bony or dense soft tissue environment. Images are produced using contrast
medium by subtracting a 'pre-contrast image' or the mask from later images, once the contrast
medium has been introduced into a structure. Hence the term 'digital subtraction angiography'.
DSA is primarily used to image blood vessels.
Catheter
A tube inserted into the body into a vessel, cavity or organ for injection or drainage purposes.
Iodinated Contrast Media
(ICM) or ‘contrast’: A liquid dye containing Iodine used mainly by injection to enhance vessels and
organs, it is visible under X-ray.
ISBAR
Clinical Handover mnemonic for Introduction/Identification, Situation, Background, Assessment,
Recommendations.
Fluoroscopy
An x ray procedure visualising internal organs in motion
4. ROLES AND RESPONSIBILITIES:
4.1. Scope:
This guideline applies to all Registered Nurses, Radiographers and Medical Officers (MO) working
within Diagnostics.
4.2. Roles:
Interventional Radiologist: Responsible for ensuring the patient is medically fit for the procedure,
has a clinical need for the procedure and is without contraindications; obtaining informed consent
and carrying out the procedure. Prescribing relevant medication; carrying out appropriate
documentation, including post procedure instructions and discharge letter (for outpatients).
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Ultimately responsible for the medical care and safety of the patient, the Radiologist will provide
the patient with suitable education about the procedure and aftercare.
Scrub Nurse: Responsible for being the sterile scrub nurse and preparing the sterile equipment,
maintaining a sterile environment, counting of instruments and equipment and skin preparation for
the patient. Assisting the MO with the procedure and safe/appropriate disposal of equipment
including sharps. The scrub nurse is responsible for signing the operating record, scout sheet and
saving data in the electronic medical system.
Scout Nurse: Responsible for the nursing care of the patient; assisting the scrub nurse with the
sterile set up, equipment count and opening sterile equipment. Observing the patient; monitoring
vital signs and accurately documenting the procedure. Responsible for providing the patient with
suitable education about the procedure and aftercare.
Sedation Nurse: If sedation is ordered by the MO the sedation nurse is responsible for ensuring
that there is a valid prescription; obtaining the sedation drugs with another RN or MO, checking
them and administering them under close medical supervision as prescribed. All sedation drugs
administered should be documented accurately and disposed of appropriately. This nurse is also
responsible for initiating oxygen therapy as requested by the MO for sedation purposes.
5. PROCESS:
5.1. Equipment:
As per Medical Imaging Nursing Procedure Setups – Interventional Radiology (DSA)
 Venous Sampling – Adrenal 1 pack

Radiology Kit 2-3 packs

Sterile Gowns (Doctor/s + Nurse) 3-4 pairs

Sterile Gloves (Doctor/s + Nurse) 1 bag

Heparinised Saline
o
1L 0.9% Sodium Chloride Added- 5000 IU Heparin Injection

2 tubes Povidone-Iodine Solution (10% w/v Cutaneous Solution)

1 amp Lignocaine Injection 1% ( 50mg/5ml)

1 pc 10 ml slip syringe (Venous Puncture)

1 pc OpSite Post-Op 6.5 x 5cm (Dressing)

1 bottle 75 mL Ultravist 300

1 pc 5FR sheath or Dr's choice

1 pc MKB1 catheter with side holes

1 pc C1/C2 Catheter with side holes Dr's choice

1 pc 150 cm (0.035") ANGLED Guide Wire Terumo
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
1 pc Ultrasound probe cover 6-7 pcs 10 ml lock syringes

12 pcs 21G green needles

4 kidneydish Ice Container

Pathology Specimen Tubes (Properly labelled – see guide below)
o
At least 2. 5mls of blood each tube

IVC Above Adrenal Veins – 1 sample (GOLD top tube)

IVC Below Adrenal Veins –1 sample (GOLD top tube)

Right Adrenal Vein – 1 sample (GOLD top tube)

Left Adrenal Vein – 1 sample (GOLD top tube)

4 pc specimen bag

4 pc Pathology request form with site of collection properly labelled

1 pc Small trolley (Sterile Gowns / Gloves)

1 pc Long Trolley (Procedure Set-up)
N.B. Use ONLY referrer request pathology forms and samples
5.2. Preparation:
Environment: The room will be a clean environment suitable for sterile procedures, all equipment
required for the procedure and patient monitoring will be available.
Patient: The Patient will undergo informed consent by the Radiologist or referring team. The patient
will be admitted through DPC (outpatients), patient history, medications, allergies and fasting
status will be documented. Site preparation performed. Baseline vital signs will be recorded. Blood
sugar to be recorded for diabetic patients. All relevant paperwork will accompany the patient for the
duration of their stay.
Attire: The patient must undress fully and wear a patient gown and disposable underwear, long hair
must be tied back and a surgical hat must be worn. Jewellery and dentures may be worn, if
documented.
Fasting: Fasting not required unless patients requiring IV sedation in which case must fast for 6
hours.
Pathology: Coagulation profile, creatinine level, electrolytes, 24 hour urine sodium level
(>100mmol/24 hours)
Medications: Continue current medications, unless otherwise advised.
The following medications will be withheld prior to the procedure:
 Warfarin: Withhold for 5 days under medical instruction

Clexane: Withhold s/c injections for 24 hours under medical instruction

Heparin: Withhold s/c injections for 24 hours under medical instruction

Clopidogrel (Plavix): Withhold for 5 days under medical instruction
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
Aspirin (Cartia): Withhold for 5 days under medical instruction

Metformin: Withhold in renal impairment for 48 hours post procedure under medical
instruction
5.3. Procedure Steps:
Diagnostics staff will verify Patient’s identity and match details against documentation, as the
Patient enters the room. Oxygen saturations and heart rate will be monitored and recorded
throughout; further vital signs monitoring as required. Hair will be removed with clippers if
necessary.
Maintaining compliance with SVH Manual Handling policy and procedure, patient is positioned onto
X-ray table in the supine position and connected to monitoring equipment. Patient’s skin is
washed with antiseptic prep solution and a sterile drape is positioned over puncture site. Timeout
will be confirmed with the senior Radiologist and the team when the MO enters the room. Local
anaesthetic is administered to anaesthetise the skin at the puncture site. A 5ml luer slip syringe
fitted with entry needle and filled with normal saline (to visualise venous access), is introduced into
femoral vein and then a wire is passed through the entry needle. Needle is removed and a
catheter/sheath is inserted over the wire and positioned in the adrenal vein. Wire is removed and
digital fluoroscopic images are obtained using the recommended contrast agent. Radiologist
obtains site specific blood samples for analysis in quick succession. Scout staff label each sample
specifically with site, time and collection order, as well as correct patient details. Each sample is
analysed for aldosterone and cortisol. Send samples directly to biochemistry. Once all images and
samples have been obtained, the Radiologist removes the catheter and applies pressure at the
puncture site until haemostasis is achieved and a clear occlusive dressing applied.
Notes:Correct labeling of all samples with date, time, site is vital for interpreting results.
5.4. Disposal of Waste/Equipment:
Waste will be disposed of according to local policy; sharps and instruments will be disposed of
appropriately by the scrub nurse or Radiologist.
5.5. Post Procedure Patient Management:
Post procedure, the patient will be transferred to Recovery Room 2 in a bed. Handover will be
given as per ISBAR Clinical Handover guidelines. The patient will have vital signs, puncture site
and limb observations monitored, as per Radiologist’s instructions. The wound will again be
checked prior to discharge, and the patient will be offered food and drink. The patient can be
discharged, or transferred to a ward bed, if the wound is not actively bleeding, when vital signs are
within normal limits and the patient feels well and not in pain. The MO will provide a discharge
letter and a script for analgesia, if appropriate.
5.6. Documentation:
Following ISBAR guidelines, DPC nursing pathway booklet will be completed on the appropriate
pages, including pre-op checklist, observation chart, a green Timeout sheet, a duplicated scout
sheet and an operation record will be completed. The procedure will be recorded in the
Interventional Radiology (DSA) and MO’s diaries and an electronic scout sheet will be recorded in
TMS. Both RN and MO must complete documentation within the pathway for outpatients or in the
clinical notes for an inpatient and ensure all medications are prescribed accordingly.
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6. Compliance:
Compliance will be monitored biannually by observational and patient medical records auditing of a
randomly selected group of patients that have had Adrenal Venous Sampling. The audit will assess
patient outcomes and best practice.
Patient Outcomes
The patient will have received optimal care before, during and after Adrenal Venous Sampling, with
no adverse results experienced.
Best Practice
 All documentation contains

Patient name, date of birth, MRN
 Evidence of informed consent

Patient’s pain assessment and management is documented

Correct Patient, Correct Procedure, Correct Site compliance
 Sterile technique competency

Adherence to Infection Control Policy
 Radiation safety
Results will be monitored and discussed by Quality Improvement Committee, Diagnostic Services
Department.
Expected KPI for compliance: 100% of audits will be compliant with clinical competency and
accuracy of documentation.
7. References: Supporting Evidence:
MINA (Medical Imaging Nurses Associations Australia) 2008 Procedural Document for Venous Sampling Adrenal. Available at http://www.minanational.com accessed July 14th 2011
NSW Department of Health (2007) Correct Patient, Correct Procedure and Correct Site. Sydney, Australia.
(PD2007_079)
ANZCA (Australian and New Zealand College of Anaesthetists) 2010 PS9 – Guidelines on Sedation and/or
Analgesia for Diagnostic and Interventional
Medical, Dental or Surgical Procedures. Available at http://www.anzca.edu.au accessed August 10th 2011.
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CLONIDINE SUPPRESSION TEST
Clonidine is an adrenergic agonist, which is useful in the diagnosis of Phaeochromocytoma.
Patient is to fast overnight and to be supine during the test.
Patient is not to be left & to remain on bed rest throughout the test. After 0 level patient can eat &
drink.
Check patient’s BP ½ hourly during the test and continue to monitor until BP returns to normal.






Site butterfly for commencement of IV fluids – normal saline. Fluids to commence at 0 level to
keep vein patent – run fluids slowly until BP returns to normal.
Clonidine 300 mcg to be given orally.
Collect blood for Catecholamines at:
- - 30 mins
- 0 level after patient rested for 30 mins
- 60 mins
- 120 mins
- 180 mins
Catecholamines are to be collected 10 mL or minimum 8 mL in special tube. These tubes are
available at pathology collection centre on each side within the area health.
Deliver on ice to Clinical Chemistry ext 29075.
To be centrifuged immediately.
SIDE EFFECTS




Drowsiness.
Fall in blood pressure.
Drugs for Reversal, if Hypertensive give Regitine 2mg initially; if Hypotensive give Atropine
500 mg slowly.
Patient is not to drive home by himself, transport need to be organised prior to admission.
REFERENCE RANGE
Catecholamines suppress in normal subjects but not in patients with phaeochromocytoma.
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Bone Biopsy Protocol including dynamic histomorphometry
Indications:







Diagnosis of osteomalacia and other bone mineralization disorders; e.g. renal
osteodystrophy (CKD-MBD).
To look for an underlying cause for an unexplained fracture and parameters of bone
turnover in these unusual cases or compliance to therapy or extent of bone formation or
resorption under basal conditions or following treatment
Evaluate bone pain or tenderness (non dynamic)
Investigate an abnormality seen on X-ray(non dynamic)
Determine if a bone tumor is malignant (cancerous) or benign(non dynamic)
Determine the cause of an unexplained infection or inflammation(non dynamic)
Recommended by an endocrinologist
Bone biopsy labelling medication prior to procedure
1. Tetracycline 500 mg bd (preferred medication if available) or Ledermycin 150 mg tds or
Vibramycin 100 mg tds or Doxycycline 100 mg tds for 2 days
2. Do not take the medication for the next 10 days
3. Tetracycline 500 mg bd (preferred medication if available) or Ledermycin 150 mg tds or
Vibramycin 100 mg tds or Doxycycline 100 mg tds for 2 days
4. Bone Biopsy scheduled for 3-5 days after the second label
Pharmacy may need to be approached to provide a supply of this medication for this procedure.
The medication is usually administered after meals to avoid gastrointestinal discomfort.
Dairy products and aluminium-containing antacids will interfere with adequate absorption and
binding of medication. Patients should be instructed to avoid milk and dairy products, as well as
calcium containing binders for 2 hours before and after medication. The patient should cease
Aspirin 7 days before the procedure. Warfarinised patients would benefit from transfer to Clexane
ceased on the day of the procedure and recommenced soon after. Advice from haematologists
about anticoagulation should be sought.
Avoiding sun exposure to prevent skin photosensitivity.
Procedure:


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The SAMPLE/BIOPSY specimen is to remain UNDECALCIFIED. It has to be done in
Radiology by Dr. Ryan Rudolf (Tel: 93820300) who will obtain the best core biopsy from
the sacral crest. This is PREFERRED.
Alternatively if Dr Rudolph is NOT available the haematology registrars (Page
44291/44237) can assist with obtaining a bone trephine of suitable quality to ensure valid
analysis BUT this risks the sample being handled in the same way as routine trephine
biopsy specimens. This is a DISASTER as routine trephines are decalcified and the effort
of labelling the specimen is lost. The endocrine registrar or CNS endocrinology needs to be
on site at the time of sample collection to ensure satisfactory sample handling under these
circumstances.
Please liaise with our endocrine nurse (Fun Chan) to inform her the date of the procedure
for specimen handling.
CHEMAI - Dynamic Function Protocol - 03
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
Bone Biopsies are done so rarely in this department that the Endocrine registrar is usually
not adequately skilled to ensure accurate, adequate and valid sampling
Fixative to store the bone sample
Preferred fixative
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Normal Saline --------------------900mls
37% formaldehyde ---------------100mls
Sodium barbital ---------------approx 4 g
Wear protective clothing, eye goggles and gloves for the following procedures.
Add the formaldehyde to the normal saline. Mix
Add sodium barbital gradually until the pH of the solution is 7.0
Store the solution at 4 C
Alternative fixative
10% neutral buffered formalin
Transport of the specimen fixation
The specimen needs to be stored at 4ºC during transportation.
The bone biopsy should be placed into one of the above fixative solutions immediately after it is
harvested and delivered through Courier at once to:
Department of Anatomical Pathology
SEALS-Kogarah
4th floor, clinical services building
St George Public Hospital
Kogarah NSW 2217
Please contact the above department 91133417 or 91133420 at the time of collection of the
specimen and the time to be expected to arrive.
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APPENDIX 1
SYMPTOMS OF HYPOGLYCAEMIA
TREMBLING / SHAKING
SWEATY / CLAMMY
PALLOR
HUNGER
IRRITABILITY
PALPITATIONS
HEADACHE
TINGLING OF FINGERS AND LIPS
DISTURBANCE OF CONCENTRATION
ABNORMAL SPEECH
BLURRED VISION
AGGRESSIVE BEHAVIOUR
CONFUSION
LOSS OF CONSCIOUSNESS
FITTING
This is not an exclusive list individual patients may experience symptoms that never occur in
another.
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APPENDIX 2
CHEMAI - Dynamic Function Protocol - 03
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APPENDIX 3
CHEMAI - Dynamic Function Protocol - 03
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CHEMAI - Dynamic Function Protocol - 03
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Authorised by: C. White & R. Horvath
Released: 18.11.2015
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