Weill B J, Heavey A C, Kelley V E, Schur P H
J Immunol Methods. 1983 Feb 25;57(1-3):327-40. doi: 10.1016/0022-1759(83)90093-5.
By studying antinuclear antibody production at the cellular level, we can better understand the problems of immunoregulation in individuals with systemic lupus erythematosus. To date, the use of hemolytic plaque assays to detect B cells secreting antinuclear antibodies has been hampered by an inability to achieve reliable coating of red cells by nuclear antigens. Because the chromic chloride technique has proved ineffective for coupling nucleic acids and/or nuclear antigens to sheep red blood cells (SRBC) in our laboratory, we have developed a method of coupling SS DNA, DS DNA, poly(I).poly(C), Sm, and ENA to red cells pretreated with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (ECDI) and poly(L-lysine) (PLL). Coated red cells were agglutinated by specific antisera and not by normal sera and were then used in hemolytic plaque assays to detect antinuclear plaque-forming cells (PFC) in spleens from various strains of mice with lupus-like syndromes. PFC specific for SS DNA, DS DNA, poly(I).poly(C), Sm, and ENA were found in MRL/lpr and NZB x W mice, and the number of anti-SS DNA and anti-DS DNA PFC correlated with the age of the animals. Indirect (IgG) PFC specific for nuclear antigens increased dramatically in female NZB x W mice between 11 and 13 months, a time when more than 50% of the animals usually die. Preliminary studies have shown that PFC specific for nuclear antigens can be detected in peripheral blood from patients with lupus erythematosus. Pretreatment of sheep red cells with ECDI and PLL thus allowed the coupling of selected nuclear antigens to these cells and provided the first demonstration of IgM and IgG PFC specific for a variety of nuclear antigens.
通过在细胞水平研究抗核抗体的产生，我们能够更好地理解系统性红斑狼疮患者的免疫调节问题。迄今为止，由于核抗原无法可靠地包被红细胞，利用溶血空斑试验检测分泌抗核抗体的B细胞受到了阻碍。因为在我们实验室中，氯化铬技术已被证明在将核酸和/或核抗原与绵羊红细胞（SRBC）偶联方面无效，所以我们开发了一种将单链DNA（SS DNA）、双链DNA（DS DNA）、聚肌苷酸-聚胞苷酸（poly(I).poly(C)）、Sm和可提取核抗原（ENA）与经1-乙基-3-(3-二甲基氨基丙基)碳二亚胺（ECDI）和聚-L-赖氨酸（PLL）预处理的红细胞偶联的方法。包被的红细胞被特异性抗血清凝集，而不被正常血清凝集，然后用于溶血空斑试验，以检测患有狼疮样综合征的各种品系小鼠脾脏中的抗核空斑形成细胞（PFC）。在MRL/lpr和NZB×W小鼠中发现了针对SS DNA、DS DNA、poly(I).poly(C)、Sm和ENA的PFC，并且抗SS DNA和抗DS DNA PFC的数量与动物年龄相关。在11至13个月大的雌性NZB×W小鼠中，针对核抗原的间接（IgG）PFC显著增加，此时超过50%的动物通常会死亡。初步研究表明，系统性红斑狼疮患者外周血中可检测到针对核抗原的PFC。因此，用ECDI和PLL对绵羊红细胞进行预处理，使得选定的核抗原能够与这些细胞偶联，并首次证明了针对多种核抗原的IgM和IgG PFC。
