由糖蛋白B（GPB）.s肽免疫产生的鼠单克隆抗-s和其他抗糖蛋白B抗体。

Murine monoclonal anti-s and other anti-glycophorin B antibodies resulting from immunizations with a GPB.s peptide.

作者信息

Halverson Gregory R, Tossas Edith, Velliquette Randall W, Lobo Cheryl, Reid Marion E, Frame Tom, Castilho Lilian, Lee Agnes H, Storry Jill R, Grodecka Magdalena, Waśniowska Kazimiera, Duk Maria, Lisowska Elwira

机构信息

Laboratoryof Immunochemistry, New York Blood Center, New York, New York 10065, USA.

出版信息

Transfusion. 2009 Mar;49(3):485-94. doi: 10.1111/j.1537-2995.2008.02004.x. Epub 2008 Nov 25.

DOI:10.1111/j.1537-2995.2008.02004.x

PMID:19040495

Abstract

BACKGROUND

The blood group antigens S and s are defined by amino acids Met or Thr at position 29, respectively, on glycophorin B (GPB). Commercial anti-s reagents are expensive to produce because of the scarcity of human anti-s serum. Our aim was to develop hybridoma cell lines that secrete reagent-grade anti-s monoclonal antibodies (MoAbs) to supplement the supply of human anti-s reagents.

STUDY DESIGN AND METHODS

Mice were immunized with the GPB(s) peptide sequence TKSTISSQTNGETGQLVHRF. Hybridomas were produced by fusing mouse splenocytes with mouse myeloma cells (X63.Ag8.653). Screening for antibody production was done on microtiter plates by hemagglutination. Characterization of the MoAbs was done by hemagglutination, immunoblotting, and epitope mapping.

RESULTS

Eight immunoglobulin G MoAbs were identified. Five antibodies are specific by hemagglutination for s and two MoAbs, when diluted, are anti-S-like, but additional analyses shows a broad range of reactivity for GPB. Typing red blood cells (RBCs) for s from 35 donors was concordant with molecular analyses as were tests on RBCs with a positive direct antiglobulin test (DAT) from 15 patients. The anti-s MoAbs are most reactive with peptides containing the (31)QLVHRF(36) motif, with (29)Thr. By Pepscan analyses, the anti-S-like MoAbs reacted within the same regions as did anti-s, but independently of (29)Met. One antibody was defined serologically as anti-U; however, its epitope was identified as (21)ISSQT(25), a sequence common for both GPA and GPB.

CONCLUSION

In addition to their value as typing reagents, these MoAbs can be used to phenotype RBCs with a positive DAT without pre-test chemical modification.

摘要

背景

血型抗原S和s分别由血型糖蛋白B（GPB）上第29位氨基酸甲硫氨酸（Met）或苏氨酸（Thr）定义。由于人抗s血清稀缺，商用抗s试剂生产成本高昂。我们的目标是开发能分泌试剂级抗s单克隆抗体（MoAb）的杂交瘤细胞系，以补充人抗s试剂的供应。

研究设计与方法

用GPB（s）肽序列TKSTISSQTNGETGQLVHRF免疫小鼠。通过将小鼠脾细胞与小鼠骨髓瘤细胞（X63.Ag8.653）融合产生杂交瘤。通过血凝试验在微量滴定板上筛选抗体产生情况。通过血凝试验、免疫印迹和表位作图对单克隆抗体进行鉴定。

结果

鉴定出8种免疫球蛋白G单克隆抗体。5种抗体通过血凝试验对s具有特异性，2种单克隆抗体稀释后具有抗S样反应，但进一步分析显示其对GPB具有广泛的反应性。对35名供者的红细胞进行s分型，结果与分子分析一致，对15例直接抗球蛋白试验（DAT）阳性患者的红细胞检测结果也一致。抗s单克隆抗体与含有（31）QLVHRF（36）基序及（29）Thr的肽反应最强。通过肽扫描分析，抗S样单克隆抗体与抗s在相同区域反应，但与（29）Met无关。一种抗体血清学上定义为抗U；然而，其表位鉴定为（21）ISSQT（25），这是血型糖蛋白A（GPA）和GPB共有的序列。