Kleine T O, Damm T
Referenzlabor für Liquordiagnostik, Abteilung Klinische Chemie und Molekulare Diagnostik, Deutschhausstrasse 12, 35033, Marburg, Germany.
Anal Bioanal Chem. 2003 Apr;375(8):1000-5. doi: 10.1007/s00216-003-1809-1. Epub 2003 Mar 28.
Semi-automated electrophoretic procedures in the PhastSystem (Amersham Pharmacia Biotech) with micro polyacrylamide gels (PAGs) and SDS-PAG gradients were modified to analyze IgG in human cerebrospinal fluid (CSF) and matched serum samples with respect to the molecular IgG structure L-H-H-L. Isoelectric focusing (IEF) with specific immunofixation detected discrete IgG bands in CSF standing out against a polyclonal and monoclonal background pattern in CSF and serum; they were denoted oligoclonal bands (IgG OBs) (OB assay positive) indicating IgG synthesis in the central nervous system (CNS) of patients with subacute and chronic processes of inflammatory CNS disorders; assay was negative with identical (mirror) bands in CSF and serum for other CNS processes. IgG OBs were specified as lambda (kappa) IgG subfractions, precipitated with the anti-light (L) chains lambda (kappa) and anti-heavy (H) chain fragments (Fd, Fc, C(H)2) as well as with anti-F(ab')(2), and as duplex IgGs with kappa and lambda OBs at the same pI. With SDS-PAG gradient electrophoresis and specific immunofixation more than six IgG fractions were detected and classed according to apparent molecular weights of a S-sulfonated human IgG standard; they were characterized with the monospecific antibodies against the L and H chain fragments as 25, 50, 75, 100, 125 and 150 kD fractions containing combinations of L and H chains as well as mixtures of both L and H chain fragments of varying dimensions. Generally, this molecular IgG heterogeneity could not be connected with the IgG OB heterogeneity revealed by IEF; but single OBs in the strongly alkaline pH region of PAG may correspond to H fragments with basic pI. Nevertheless, evidence for the existence of both free L chains and the free H chain were revealed as specific OBs with IEF and with the anti-L and anti-H antibodies in the 25 kD and 50 kD fractions, respectively, of CSF samples of six patients with diverse CNS diseases. Further experiments are needed to elicit the origin of the molecular IgG heterogeneity during the immune response of subacute and chronic inflammatory processes in human CNS.
对PhastSystem（安玛西亚 Pharmacia 生物技术公司）中采用微型聚丙烯酰胺凝胶（PAG）和SDS - PAG梯度的半自动电泳程序进行了改进，以分析人脑脊液（CSF）和匹配血清样本中IgG的分子结构L - H - H - L。通过特异性免疫固定的等电聚焦（IEF）在CSF中检测到离散的IgG条带，在CSF和血清的多克隆和单克隆背景模式下显得突出；这些条带被称为寡克隆条带（IgG OBs）（OB检测阳性），表明患有亚急性和慢性炎症性中枢神经系统（CNS）疾病的患者中枢神经系统（CNS）中有IgG合成；对于其他CNS疾病，CSF和血清中相同（镜像）条带的检测为阴性。IgG OBs被指定为λ（κ）IgG亚组分，用抗轻链（L）λ（κ）和抗重链（H）片段（Fd、Fc、C(H)2）以及抗F(ab')(2)沉淀，并且在相同pI处作为具有κ和λ OBs的双链IgG。通过SDS - PAG梯度电泳和特异性免疫固定检测到六个以上的IgG组分，并根据S - 磺化人IgG标准品的表观分子量进行分类；它们用针对L和H链片段的单特异性抗体表征为25、50、75、100、125和150 kD组分，包含L和H链的组合以及不同尺寸的L和H链片段的混合物。一般来说，这种分子IgG异质性与IEF揭示的IgG OB异质性无关；但在PAG强碱性pH区域的单个OBs可能对应于具有碱性pI的H片段。然而，在六名患有不同CNS疾病的患者的CSF样本中，分别通过IEF以及25 kD和50 kD组分中的抗L和抗H抗体揭示了游离L链和游离H链存在的证据。需要进一步的实验来探究人CNS亚急性和慢性炎症过程免疫反应期间分子IgG异质性的起源。
