van de Corput L, Chaux P, van der Meijden E D, De Plaen E, Frederik Falkenburg J H, van der Bruggen P
Department of Hematology, Leiden University Medical Center, Leiden, The Netherlands.
Leukemia. 2005 Feb;19(2):279-85. doi: 10.1038/sj.leu.2403583.
In patients with hematological malignancies receiving HLA-matched stem cell transplantation, T cells specific for minor histocompatibility antigens play a major role in graft rejection, induction of graft-versus-host disease and beneficial graft-versus-leukemia reactivity. Several human minor histocompatibility antigens recognized by T cells have been identified, but only two are presented by HLA class II molecules. In search of an efficient approach to identify antigenic peptides processed through the HLA class II pathway, we constructed a cDNA library in bacteria that were induced to express proteins. Bacteria were opsonized with complement to enforce receptor-mediated uptake by Epstein-Barr virus immortalized B cells that were subsequently used as antigen-presenting cells. This approach was validated with an HLA class II-restricted antigen encoded by gene DBY. We were able to identify bacteria expressing DBY diluted into a 300-fold excess of bacteria expressing a nonrelevant gene. Screening of a bacterial library using a DBY-specific CD4 T cell clone resulted in the isolation of several DBY cDNAs. We propose this strategy for a rapid identification of HLA class II-restricted antigenic peptides recognized by CD4 T cells.
在接受人类白细胞抗原（HLA）匹配的干细胞移植的血液系统恶性肿瘤患者中，针对次要组织相容性抗原的T细胞在移植物排斥、移植物抗宿主病的诱导以及有益的移植物抗白血病反应中起主要作用。已经鉴定出几种被T细胞识别的人类次要组织相容性抗原，但只有两种由HLA II类分子呈递。为了寻找一种有效的方法来鉴定通过HLA II类途径加工的抗原肽，我们在诱导表达蛋白质的细菌中构建了一个cDNA文库。用补体调理细菌，以促使其被爱泼斯坦-巴尔病毒永生化的B细胞通过受体介导摄取，随后将这些B细胞用作抗原呈递细胞。该方法通过由DBY基因编码的一种HLA II类限制性抗原得到验证。我们能够从稀释在过量300倍表达无关基因的细菌中的细菌中鉴定出表达DBY的细菌。使用DBY特异性CD4 T细胞克隆筛选细菌文库，结果分离出了几种DBY cDNA。我们提出这种策略用于快速鉴定被CD4 T细胞识别的HLA II类限制性抗原肽。
