Application Of Microarray Technology To Research The Status Of Her-2 Gene Amplification And Protein Expression In Breast Cancer

Objective: To use the tissue microarray (TMA) technology. Tissue microarray group (TMAG) and nucleus microarray (NMA) can be made successfully by paraffin embedded tissues. To compare HER-2 status of fluorescence in situ hybridization (FISH) and protein expression of immunohistochemistry (IHC) in breast cancer. Research the relationship in two methods and Clinical significance.Materials and Methods:MaterialCollected 248 cases of breast cancer paraffin embedded tissues from the General Military Hospital of Shenyang during June 2006 ~ August 2008.Making of TMAG and IHCThe TMA sections from different sources will be arranged in the same glass slice. At the same time narrowed the distance between adjacent cores to increase the number of chips. The TMAG was made successfully by paraffin embedded tissues. Use the S-P method of IHC. IHC positive shows color on the cell membrane. Immunohistochemistry results reference to the U.S Food and Drug Administration (FDA) standards: the color of cell membrane were observed for the positive (-~+++) through the light microscope. Judge the outcome of IHC+++ was positive and IHC-~++ was negative.Making of NMA and FISHDeparaffinage and digestion after Nuclei extracted from thick paraffin embedded tissues in order to obtain desired the nuclei. A blank recipient paraffin block with a lot of holes was constructed and sectioned to make the mold for the NMA. Prepared nuclei were injected into the holes of the paraffin mold. The slides were dried and dewaxed and nuclei arrays were made. FISH Probe Kit consists of two labeled DNA probes. The entire HER-2 gene is labeled in SpectrumOrange. The CEP 17 probe is labeled in SpectrumGreen. Results on enumeration of 20 interphase nuclei from tumor cells per target are reported as the ratio of average HER-2/neu copy number to that of CEP 17. Our clinical study found that specimens with amplify- cation showed SpectrumOrange and SpectrumGreen signal ratio of greater than or equal to 2.0; normal specimens showed a ratio of less than 2.0. Results at or near the cut-off point (1.8-2.2) should be interpreted with caution.Statistical methodsThe X~2 test was used to examine relationships between variables. Analyses were carried out using SPSS 11.5 software.Results:Results Detected by TMAG and NMAWe have used the TMAG/NMA technology. 2560 chips from different sources will be arranged in the same glass slice, Even if four chips taken from each of the samples. TMAG was made successfully for testing of the IHC. NMA was made successfully by paraffin embedded tissues. Prepared nuclei were injected into the hole of the paraffin mold. NMA was made successfully for testing of the FISH. Morphology and structure of nucleus without impurities was perfect.Relationship between the results of IHC and FISHThere were 90 cases, and 49 cases, and 37 cases and 72 cases from IHC- to IHC +++ in IHC staining results. 172 cases were FISH-negative and 76 cases were FISH-positive. The coincident rate of two results was 87.9 %, therefore there was a significant positive correlation between the two techniques. No significant correlation between the FISH and IHC++. Four cases of chromosome 17 monosomy were found in IHC++/FISH-positive. It was 5.26% in all the FISH-positive. Five cases of chromosome 17 polysomy were found in IHC+++/ FISH-negative. It was 2.9% in all the FISH- negative. Conclusions1. TMAG and NMA can be applied to detection of IHC and FISH in breast cancer. It is high efficient, good consistency and more reliable.2. The results of IHC and FISH showed a good coincidence when IHC-~+ and IHC+++ in the mammary cancer tissues.3. It is necessary to evaluate HER-2 gene amplification and distinguish chromosome 17 monosomy through the FISH method when IHC++. Avoid unnecessary targeted drug therapy.Innovation of this study1. TMAG and NMA can be applied to detection of IHC and FISH in breast cancer. To expand the detection means to the breast cancer.2. Proposed FISH testing is necessary to distinguish chromosome 17 monosomy in order to avoid unnecessary treatment when IHC++。Shortcomings of the researchLong term follow-up data of the patients are in shortage so that more meaningful conclusions can't be summarized.