In-cell assembly of single-chain Fv from peripheral blood lymphocytes from vitiligo patients
- VernacularTitle:In-cellPCR技术扩增白癜风自身抗体基因
- Author:
Yanli TIAN
;
Chunying LI
;
Gang WANG
;
Tianwen GAO
;
Tao LU
;
Miao LI
- Publication Type:Journal Article
- Keywords:
Vitiligo;
Polymerase chain reaction;
Auto antibodies
- From:
Chinese Journal of Dermatology
1995;0(01):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To obtain in situ pairing of the variable region genes of the immunoglobulin heavy and light chains of B cells from vitiligo patients. Methods Fifty vitiligo patients were screened by a live melanocyte enzyme-linked immunoabsorbent assay. The sera from seven patients were proved to be strongly positive. Then CD19+ B cells were isolated from the peripheral blood lymphocytes of the 7 patients, then fixed in 10% buffered formaldehyde and permeabilised by Proteinase K. The mRNA was amplified within the cells by reverse transcription-polymerase chain reaction (RT-PCR) with specific primers. The immunoglobulin VH and VL DNA assembled within the same cells using the Cre/loxP system. Nested primers were designed to amplify the known major human VH and VL gene familes. Result A unique 800bp band was obtained corresponding in size to single chain Fv fragments. Conclusion The in situ pairing of the variable region genes of the immunoglobulin heavy and light chains of B cells is obtained from vitiligo patients by in-cell PCR.
