Quantitative Assessment of HER-2/Neu Protein Concentration in Breast Cancer by Enzyme-Linked Immunosorbent Assay

2003 ◽  
Vol 18 (1) ◽  
pp. 13-20 ◽  
Author(s):  
V. Müller ◽  
C. Thomssen ◽  
C. Karakas ◽  
I. Eustermann ◽  
J. Ramirez Porras ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Gene Amplification ◽  
Immunosorbent Assay ◽  
Immunohistochemical Staining ◽  
Protein Concentration ◽  
Tumor Tissue ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cancer Tissue ◽  
Tissue Extracts ◽  
Her 2

Purpose The HER-2/neu protein (p185) has become a promising target for antibody therapy in breast cancer. We tested the feasibility of a quantitative approach for HER-2/neu testing based on the analysis of tumor tissue extracts by an enzyme-linked immunosorbent assay (ELISA). Experimental design Tumor tissue extracts of primary human breast cancers (n=124) were prepared using a triton-based buffer. HER-2/neu concentration was quantified by ELISA. Paraffin-embedded tissue sections of the same tumors were analyzed by immunohistochemical staining applying the monoclonal HER-2/neu antibody TAB 250 (n=124) and by chromogenic in situ hybridization (CISH) (n=73). Results Concentrations of p185 in tissue extracts determined by ELISA varied from 1 to 927 ng per mg protein with a median of 25 ng/mg protein, whereas normal breast tissue showed values from 0.4 to 5.5 ng/mg with a median of 2.2 ng/mg (p<0.0001, Mann-Whitney U test). A significant correlation between p185 concentration and immunohistochemical staining was observed (p<0.0001, Kruskal-Wallis test). In addition, p185 concentration measured by ELISA was correlated with the degree of HER-2/neu gene amplification determined by CISH. HER-2/neu-amplified tumors had significantly higher p185 concentrations (median value 181 ng/mg protein) than non-amplified tumors (median value 20 ng/mg; p<0.0001, Mann-Whitney U test). Conclusions ELISA-based measurement of HER-2/neu protein concentration in breast cancer tissue extracts is feasible and provides quantitative results for p185 protein concentrations that correlate closely with HER-2/neu immunoscore and gene amplification.

HER-2/neu Analysis in Archival Tissue Samples of Human Breast Cancer: Comparison of Immunohistochemistry and Fluorescence In Situ Hybridization

2001 ◽  
Vol 19 (2) ◽  
pp. 354-363 ◽  
Author(s):  
Annette Lebeau ◽  
Daniela Deimling ◽  
Christine Kaltz ◽  
Andrea Sendelhofert ◽  
Anette Iff ◽  
...  
Keyword(s):  
Breast Cancer ◽  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Gene Amplification ◽  
Cancer Tissue ◽  
Breast Cancers ◽  
Paraffin Sections ◽  
Tissue Samples ◽  
Her 2

PURPOSE: The objective of our study was to compare the methods used in the literature to analyze HER-2/neu status on archival breast cancer tissue. Therefore, a series of antibodies was evaluated to assess their immunohistochemical (IHC) sensitivity in correlation to gene amplification determined by fluorescence in situ hybridization (FISH). MATERIALS AND METHODS: HER-2/neu overexpression was studied on paraffin sections of 85 invasive breast cancers using a panel of five monoclonal (9G6, 3B5, CB11, TAB250, GSF-HER2) and two polyclonal antibodies (A8010, A0485) in addition to the HercepTest (DAKO, Glostrup, Denmark). HER-2/neu gene amplification was determined by FISH using a dual-color probe (PathVysion; Vysis, Stuttgart-Fasanenhof, Germany). RESULTS: HER-2/neu overexpression was demonstrated in 26% (9G6, TAB250, GSF-HER2), 27% (3B5, CB11), 33% (A8010) and 42% (A0485, HercepTest) of the tumors. FISH on paraffin sections identified gene amplification in 28% of the tumors. Strongly positive IHC results (3+) were always associated with gene amplification. Among the 16 tumors presented with weakly positive IHC results (2+) using the HercepTest, 12 (75%) lacked gene amplification. CONCLUSION: The comparison of IHC and FISH demonstrated an excellent correlation of high-level HER-2/neu overexpression (3+) with gene amplification; ie, FISH does not provide further information in these tumors. However, weakly positive IHC results (2+) obtained with the HercepTest share only a minor association with gene amplification.

Immunohistochemical Staining of Metastatic Ductal Carcinomas of the Breast by Monoclonal Antibodies used in Imaging and Therapy: A Comparative Study

1995 ◽  
Vol 10 (3) ◽  
pp. 129-135 ◽  
Author(s):  
L.P. Howell ◽  
S.J. Denardo ◽  
N.B. Levy ◽  
J. Lund ◽  
G.L. Denardo
Keyword(s):  
Breast Cancer ◽  
Monoclonal Antibodies ◽  
Immunohistochemical Staining ◽  
Tumor Tissue ◽  
Ductal Carcinoma ◽  
Immunoperoxidase Staining ◽  
Cancer Tissue ◽  
Carcinoma Of The Breast ◽  
Normal Tissues ◽  
Metastatic Lesions

Five monoclonal antibodies (MoAbs) (L6, 170H.82, 155, BrE-3 and BR96), most of which have been previously shown to target breast cancer and not normal tissues by immunoscintigraphic imaging, were evaluated for their frequency and pattern of immunohistochemical staining in 67 to 116 metastatic lesions from patients with ductal carcinoma of the breast. Immunoperoxidase staining in 75% or more of the cells occurred in 56/116 (48%) for L6, 44189 (49%) for Br, -96, 58/102 (57%) for 155, 62/99 (84%) for 170H.82, and 65.67 (97%) for BrE-3. With the first three MoAbs, an additional 6-10% of the tumors showed staining in 50-75% of tumor cells. These results illustrate that most patients with metastatic ductal carcinoma have cancer tissue in which a high percent of cells will react to several of these selected MoAbs that target different epitopes. The high expression of the MoAb targets throughout the tumor tissue makes these antibodies potential candidates to carry immunologically directed radioimmunotherapy and is an aid in selecting patients for treatment..

scholarly journals EORTC Receptor and Biomarker Study Group Report: A Sandwich Enzyme-Linked Immunosorbent Assay for Vascular Endothelial Growth Factor in Blood and Tumor Tissue Extracts

2000 ◽  
Vol 15 (2) ◽  
pp. 184-191 ◽  
Author(s):  
P.N. Span ◽  
N. Grebenchtchikov ◽  
J. Geurts-Moespot ◽  
J.R. Westphal ◽  
A.M.J. Lucassen ◽  
...  
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Growth Factor ◽  
Immunosorbent Assay ◽  
Tumor Tissue ◽  
Enzyme Linked Immunosorbent Assay ◽  
Study Group ◽  
Vascular Endothelial ◽  
Tissue Extracts ◽  
Anti Vegf ◽  
Endothelial Growth Factor

A four-antibody sandwich enzyme-linked immunosorbent assay (ELISA) for vascular endothelial growth factor (VEGF) for application in blood (serum and plasma) and tumor tissue extracts was set up within the framework of the EORTC Receptor and Biomarker Study Group (RBSG). Polyclonal antibodies against VEGF165 were raised in chickens and rabbits, and used in a previously described assay format. The assay was validated and characterized for use in serum, plasma and tumor tissue extracts. The resulting VEGF ELISA was found to be specific for VEGF165 and VEGF121, the main isoforms of VEGF. The assay showed good precision and parallelism in serial dilutions of samples. The assay was not susceptible to interference by heterophilic antibodies because avian antibodies (duck anti-chicken and chicken anti-VEGF) were used in the pre-analyte stage and mammalian antibodies (rabbit anti-VEGF and goat anti-rabbit) in the post-analyte stage. In conclusion, a sensitive, robust and specific VEGF ELISA has been developed. Research into the prognostic value of VEGF employing this assay is currently underway.

Assay of the c-erbB-2 Oncogene Encoded Protein by ELISA and Immunocytochemistry in Human Breast Cancer

1994 ◽  
Vol 31 (2) ◽  
pp. 171-173 ◽  
Author(s):  
A Nugent ◽  
J Gallagher ◽  
J Dolan ◽  
N O'Higgins ◽  
M J Duffy
Keyword(s):  
Breast Cancer ◽  
Gene Amplification ◽  
Immunosorbent Assay ◽  
Human Breast Cancer ◽  
Enzyme Linked Immunosorbent Assay ◽  
Breast Cancers ◽  
Human Breast

The c-erbB-2 gene is amplified in a wide variety of different adenocarcinomas. Generally, gene amplification correlates with increased expression of the c-erbB-2 oncoprotein. Previous assays for the c-erbB-2 oncoprotein have been qualitative or semi-quantitative. In this investigation using human breast cancers, c-erbB-2 oncoprotein levels as measured by enzyme-linked immunosorbent assay (ELISA) correlated significantly with semi-quantitation by immunocytochemistry ( r = 0·843, P<0·0001, n = 97). The cut-off point for the ELISA which gave the strongest agreement with immunocytochemistry was 15 units/μg protein. It is concluded that detection of c-erbB-2 oncoprotein by ELISA is quantitative and objective.

A PILOT STUDY OF HER-2/NEU GENE AMPLIFICATION ASSESSED BY FLUORESCENCE IN SITU HYBRIDIZATION (FISH) IN GASTRIC CANCER

2014 ◽  
pp. 15-20
Author(s):  
Van Huy Tran ◽  
Thi Minh Thi Ha ◽  
Trung Nghia Van ◽  
Viet Nhan Nguyen ◽  
Phan Tuong Quynh Le ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Cancer Patients ◽  
Gene Amplification ◽  
Cancer Tissue ◽  
Tissue Samples ◽  
Her 2 ◽  
Gastric Cancer Patients

Background: HER-2/neu is a predictive biomarker for treatment of gastric cancer using trastuzumab in combination with chemotherapy. This study aimed to evaluate the status of HER-2/neu gene amplification using fluorescence in situ hybridization (FISH) in gastric cancer. Patients and methods: thirty six gastric cancer patients were assessed HER-2/neu gene amplification by FISH using PathVysionTM HER-2 DNA Probe kit (including HER-2/neu probe and CEP-17 probe) with biopsy and surgical specimens. Results: The HER-2/neu gene amplification was observed in three cases (8.3%), the HER-2/neu gene amplification rate in Lauren’s intestinal-type and diffuse-type were 11.8% and 5.2%, respectively. Conclusion: We applied successfully FISH technique with gastric cancer tissue samples. This technique could be performed as routine test in gastric cancer in order to select patients that benefit from trastuzumab in combination with chemotherapy.

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