Download Antibodies Targeting ASGPR and PanCK for Capture and

Antibodies Targeting ASGPR and PanCK for
Capture and Identification of Circulating
Hepatocellular Carcinoma Cells
Application Note
Introduction
● Hepatocellular carcinoma is the fifth most common cancer in the world and has the highest incidence in Asia countries1.
● Biomarkers for capture and identification of circulating tumor cells (CTCs) in patients with hepatocellular carcinoma (HCC) are not
well known2.
● Although HCC cells are epithelial cells amenable to CTC capture, only about 35% of the HCC cases express EpCAM biomarker3.
● Sialoglycoprotein receptor (ASGPR), found exclusively on the surfaces of hepatocytes, binds and internalizes molecules with terminal galactose or N-acetylgalactosamine4.
● In this Application Note, we applied CytoQuest™ CR system, ASGPR and PanCK monoclonal antibodies to capture and identify
circulating HCC cells.
Materials & Methods
● Peripheral blood of HCC patient was collected in Heparin Tube (02-689-6, BD).
● 4.5 mL blood was prepared for collecting the peripheral blood mononuclear cell (PBMC) by density gradient centrifugation using
Leucosep® (163290P, Greiner Bio-One) and Histopaque®-1077 (10771, Sigma-Aldrich).
● The PBMC fraction was harvested and resuspended in Wash Medium.
● Resuspended PBMC was loaded into the CytoQuest™ CR System and HCC CTCs were captured by ASGPR (KA4573, Abnova)
immobilized CytoChipNano (U0095, Abnova).
● Immunofluorescence staining for detecting HCC CTCs were performed using PanCK, CD45 (KA4573, Abnova), DAPI as the instruction of protocol.
● Imaging was performed using Nikon Eclipse Ti-E fluorescent inverted microscope.
Results
● HCC CTC Counts: In 4.5 mL blood of HCC patient, 110 cells count as HCC CTC (PanCK+, CD45-, DAPI+).
Merged
PanCK
CD45
Nucleus
Figure 1. Representative images of HCC CTC (white arrow) and WBCs (yellow arrow) from HCC patient. HCC CTC was
detected by using immunofluorescence staining for PanCK (FITC, green), CD45 (PE, orange) and Nucleus (DAPI, blue).
Discussions
● Clinical management of HCC is mostly hampered by the lack of effective therapies and inability to monitor early metastasis and
recurrence.
● Conventional CTCs are capture and identified based on patient’s tumors and their epithelial cell derivation via EpCAM and PanCK
biomarkers.
● A new ASGPR and PanCK biomarker cocktail successfully capture and identify HCC cells using a microfluidic-based CytoQuest™
CR system.
● Therefore, HCC specific CTC detection and monitoring can provide clinically insights into treatment attempts, metastasis and
prediction of prognosis.
www.abnova.com
Abnova Corporation 9F, No. 108, Jhouzih St., Neihu, Taipei 114, Taiwan
Tel: + 886 2 8751 1888 Fax: + 886 2 6602 1218
References
1. Bosetti C, Turati F, La Vecchia C. Hepatocellular carcinoma epidemiology. Best practice & research clinical gastroenterology, 2014,
28(5):753-70.
2. Chiappini F. Circulating tumor cells measurements in hepatocellular carcinoma. International journal of hepatology, 2012,
2012:684802.
3. Yamashita T, Forgues M, Wang W, Kim JW, Ye Q, Jia H, Budhu A, Zanetti KA, Chen Y, Qin LX, Tang ZY, Wang XW. EpCAM and
alpha-fetoprotein expression defines novel prognostic subtypes of hepatocellular carcinoma. Cancer research, 2008, 68(5):145161.
4. Ise H, Nikaido T, Negishi N, Sugihara N, Suzuki F, Akaike T, Ikeda U. Effective hepatocyte transplantation using rat hepatocytes
with low asialoglycoprotein receptor expression. The American journal of pathology, 2004, 165(2):501-10.
www.abnova.com
Abnova Corporation 9F, No. 108, Jhouzih St., Neihu, Taipei 114, Taiwan
Tel: + 886 2 8751 1888 Fax: + 886 2 6602 1218