Tuaillon N, Watts R A, Isenberg D A, Muller S
Laboratoire d'Immunochimie, UPR 9021 CNRS, Institut de Biologie Moléculaire et Cellulaire, Strasbourg, France.
Mol Immunol. 1994 Mar;31(4):269-77. doi: 10.1016/0161-5890(94)90124-4.
Two human IgM lambda monoclonal antibodies (MAb) derived from the splenic lymphocytes of patients with idiopathic thrombocytopenia (Ben) and systemic lupus erythematosus (Wri) were studied. BEN-27 and WRI-170 hybridoma supernatants were screened for binding to ssDNA, dsDNA, poly (ADP-ribose), cardiolipin, histone subclasses and Klebsiella K30 cell wall antigen. Of this panel of antigens, BEN-27 and WRI-170 antibodies reacted only with histone H1. Their fine specificity was defined by direct and inhibition ELISA with synthetic peptides of the major human H1b variant. Antibody WRI-170 was shown to bind to both the N- and C-terminal peptides encompassing residues 1-16 and 204-218 of H1b whereas BEN-27 reacted only with peptide 204-218. To analyse the genetic origin of these autoantibodies, we determined the nucleotide sequence of the heavy (H) and light (L) chain variable regions of these two hybridomas. BEN-27 and WRI-170 MAbs were found to use VH1-DN1-JH4/V lambda 3-J lambda 2 and VH3-DIR2-D21/9-JH1/V lambda 2-J lambda 2 gene segment combinations respectively. Between 70 and 95% homology was demonstrated when the mRNA sequences for BEN-27 and WRI-170 were compared with published VH and V lambda germline sequences. This finding suggests that BEN-27 heavy and light chains and WRI-170 light chain use unidentified VH and V lambda germline gene segments whereas WRI-170 heavy chain derives from a VH gene segment recently identified. It is noteworthy that the CDRs of the two MAbs contain several negatively charged amino acids which are assumed to be of critical importance in antigen binding. Moreover, striking similarities are observed between BEN-27 heavy chain CDR2 and a previously described murine anti-H1 Ab heavy chain CDR2.
对源自特发性血小板减少症患者（Ben）和系统性红斑狼疮患者（Wri）脾淋巴细胞的两种人IgMλ单克隆抗体（MAb）进行了研究。对BEN - 27和WRI - 170杂交瘤细胞培养上清液进行筛选，以检测其与单链DNA、双链DNA、聚（ADP - 核糖）、心磷脂、组蛋白亚类和肺炎克雷伯菌K30细胞壁抗原的结合情况。在这组抗原中，BEN - 27和WRI - 170抗体仅与组蛋白H1发生反应。通过使用主要人H1b变体的合成肽进行直接和抑制ELISA来确定它们的精细特异性。结果显示，抗体WRI - 170与包含H1b第1 - 16位和204 - 218位残基的N端和C端肽段均能结合，而BEN - 27仅与肽段204 - 218发生反应。为了分析这些自身抗体的遗传起源，我们测定了这两种杂交瘤重链（H）和轻链（L）可变区的核苷酸序列。发现BEN - 27和WRI - 170单克隆抗体分别使用VH1 - DN1 - JH4/Vλ3 - Jλ2和VH3 - DIR2 - D21/9 - JH1/Vλ2 - Jλ2基因片段组合。当将BEN - 27和WRI - 170的mRNA序列与已发表的VH和Vλ种系序列进行比较时，发现它们之间具有70%至95%的同源性。这一发现表明，BEN - 27的重链和轻链以及WRI - 170的轻链使用了未鉴定的VH和Vλ种系基因片段，而WRI - 170的重链来源于最近鉴定出的一个VH基因片段。值得注意的是，这两种单克隆抗体的互补决定区（CDR）包含几个带负电荷的氨基酸，据推测这些氨基酸在抗原结合中至关重要。此外，在BEN - 27重链CDR2与先前描述的鼠抗H1抗体重链CDR2之间观察到显著的相似性。
