Moore T L, Dorner R W, Zuckner J
J Lab Clin Med. 1986 May;107(5):465-70.
Sera of 12 patients with rheumatoid arthritis (RA) who had positive IgM-rheumatoid factor (RF) tests were separated by use of immunoabsorbent columns (goat anti-human C3 [alpha HC3] and rabbit anti-human C1q [alpha HC1q]) and polyethylene glycol (PEG) precipitation-protein A affinity chromatography to isolate their immune complexes (IC). The isolated fractions were assayed for 19S IgM RF and 7S IgG RF by enzyme-linked immunosorbent assay (ELISA). The sera were further analyzed by preparative isoelectric focusing (IEF). The alpha HC1q and alpha HC3 columns were sequentially eluted with barbital buffer, 0.02 mol/L EDTA, 0.5 mol/L NaCl, and 1 mol/L propionic acid. All 12 patients had IgM RF and IgG RF in the EDTA fractions from both immunoabsorbent columns, but only IgM RF in the NaCl fractions. The PEG precipitation-protein A preparations were eluted with 0.5 mol/L glycine HCl and 3.5 mol/L MgCl2. All 12 patients had significant titers of 19S IgM RF (greater than or equal to 1:192) and 7S IgG RF (greater than or equal to 1:96) in the acid-eluted fraction. Analysis of the sera by preparative IEF revealed IgM RF with a polyclonal spectrotype pattern with pH of 3.0 to 10.0, but predominantly acidic proteins with isoelectric points of 4.0 to 5.5 IgG RF were found in the same restricted spectrotypic pattern. These studies demonstrated that IC can be detected in the sera of patients with RA by isolation with alpha HC1q and alpha HC3 immunoabsorbent columns and PEG precipitation-protein A affinity chromatography.(ABSTRACT TRUNCATED AT 250 WORDS)
对12例IgM类风湿因子(RF)检测呈阳性的类风湿关节炎(RA)患者的血清,采用免疫吸附柱(山羊抗人C3[αHC3]和兔抗人C1q[αHC1q])以及聚乙二醇(PEG)沉淀 - 蛋白A亲和层析法进行分离,以分离其免疫复合物(IC)。通过酶联免疫吸附测定(ELISA)对分离出的组分检测19S IgM RF和7S IgG RF。通过制备性等电聚焦(IEF)对血清作进一步分析。αHC1q和αHC3柱依次用巴比妥缓冲液、0.02mol/L乙二胺四乙酸(EDTA)、0.5mol/L氯化钠和1mol/L丙酸洗脱。所有12例患者在两种免疫吸附柱的EDTA组分中均有IgM RF和IgG RF,但在氯化钠组分中仅有IgM RF。PEG沉淀 - 蛋白A制剂用0.5mol/L甘氨酸盐酸盐和3.5mol/L氯化镁洗脱。所有12例患者在酸洗脱组分中均有显著滴度的19S IgM RF(大于或等于1:192)和7S IgG RF(大于或等于1:96)。通过制备性IEF对血清分析显示,IgM RF具有pH值为3.0至10.0的多克隆光谱型模式,但在相同的受限光谱型模式中发现主要是等电点为4.0至5.5的酸性蛋白的IgG RF。这些研究表明,通过αHC1q和αHC3免疫吸附柱分离以及PEG沉淀 - 蛋白A亲和层析法可在RA患者血清中检测到IC。(摘要截短至250字)