Identification Of Anti-hMAM And Preliminary Study Of Clinical Meaning

hMAM located on chromosome11q13, encodes93aminoacids. Its average molecularweight is10.5KD. hMAM which has specificity of breast tissue belongsto super-family of uterine protein, and it can be applied to the breast cancer earlydiagnosis, prognosis, micrometastasisthe detection, with good prospects for clinicalapplicationObjective We have got the immunogen with high purity with the method ofgenetic engineering, then Immunized mice,.After that, we obtain specific anti-hMAMmonoclonal antibody by hybridoma technology, Now on this basis, the Thermocompany Melon, Gel Method Monoclonal IgG Purification Kit is used to purifythe two anti-hMAM monoclonal antibodies, then, we identify their correspondingepitopes. Some cases are selected randomly, including primary breast cancer,benignbreast disease and normal breast tissues, breast cancermetastases, systemic non-breasttumor tissue., we analysis the sensitivity and specificity respectively using twomonoclonal antibody and compare with monoclonal antibodies and polyclonal-antibody which are foreign imports. We identify whether there is mammaglobin inprotein extracted from fresh breast cancer, benign breast lesions and normal breasttissue using monoclonal antibodies purified with the method of Western Blot.Methods1.To purify anti-hMAM antibodies Using Thermo’s Melon, GelMethod Monoclonal IgG Purification Kit and to identify epitope Corresponding to theantibody.2. To detect the sensitivity and specificity of the two antibodies using immuno-histochemical techniques and compare with the antibody import. 3. To identify expression of hMAM in protein extracted from fresh tissue of breastrelated organizations with the method of Western blot.Result1. Two anti-hMAM monoclonal antibody with higher concentrations thatis1.7g/L and2.7g/L was obtained respectively using the above-mentionedpurification method Western blotting results show that MEF521andMDA822identify hMAM43-70、56-82AA、 hMAM20-52AA-specific monoclonalantibody fragment respectively.2. Mammaglobin protein (hMAM) is expressed in tissues of breast cancer, parane-oplastic, normal breat and fibroadenoma by Western blotting which isfind two specific bands.3. Immunohistochemical staining results showed that mammaglobin is expressdpositively only in the breast related organizations. It is expressed by30%-80%ofMetastasis tissues of breast cancer, but there is no positive expression.in non-breasttissue.Conclusion: We got two anti-hMAM monoclonal antibodies and their purity wasvery good using the method which is mentioned. Compared with ab81611, we findthat MDA822and MEF521is more specificity with the method ofIHC. Sensitivity and specificity are similsr with rabbit anti-92044. Expression ofhumman mammaglobin is detaected in breast cancer, benign lesions and normalbreast tissues cases by immunohistochemistry and Western blot.