Collazo Marta E, Díaz Ana María
Department of Microbiology, School of Medicine, Medical Sciences Campus, University of Puerto Rico, PO Box 365067, San Juan, Puerto Rico 00936-5067.
P R Health Sci J. 2003 Dec;22(4):345-51.
To produce and characterize monoclonal antibodies (mAbs) against whole body extract of the dust mite Blomia tropicalis (Bt).
Bt is an important source of allergens causing allergic diseases in tropical and subtropical regions. mAbs are excellent tools for delineating cross-reactivity between Bt and other mites.
Mice were immunized with extracts of Bt. mAbs were produced by standard techniques. Hybridoma screening was performed by ELISA. Ascitic fluids were produced and partially purified by adsorption chromatography. Reactivity of mAbs against extracts of Bt and Dermatophagoides pteronyssinus (Dp) was analyzed by ELISA, immunoblots and ELISA inhibition. Also, reactivity of mAbs against the putative cysteine protease rBlo t 1 from Bt was tested.
Three IgG mAbs were obtained and partially purified. The mAbs reacted with Bt by ELISA. In immunoblots, mAbs recognized two protein components of 29 and 35 KD. Also, the mAbs reacted with Dp extracts by ELISA, and the same sized components were detected in immunoblots. In competitive ELISA, Dp extract reduced in 62% the reactivity with Bt antigens, and Bt extract produced a maximum reduction of reactivity against Dp antigens of 83%. The mAbs recognized rBlo t1. The homology between Blo t1 and the proteins recognized by the mAbs was high (90%) as the inhibition assays demonstrated.
The cross-reactivity between Dp and Bp could be related with the presence of proteins as their respective cysteine proteases. The produced mAbs have proven to facilitate the identification of antigens of Bt and to determine the possible cross-reactivity between Bt and other common mites of the acarofauna of tropical and subtropical countries where Bt is commonly found.
制备并鉴定抗热带粉尘螨（Bt）全虫提取物的单克隆抗体（mAb）。
Bt是热带和亚热带地区引起过敏性疾病的重要过敏原来源。单克隆抗体是描绘Bt与其他螨类之间交叉反应性的优秀工具。
用Bt提取物免疫小鼠。通过标准技术制备单克隆抗体。采用酶联免疫吸附测定（ELISA）进行杂交瘤筛选。制备腹水并通过吸附色谱法进行部分纯化。通过ELISA、免疫印迹和ELISA抑制分析单克隆抗体对Bt和屋尘螨（Dp）提取物的反应性。此外，还测试了单克隆抗体对来自Bt的假定半胱氨酸蛋白酶rBlo t 1的反应性。
获得了三种IgG单克隆抗体并进行了部分纯化。这些单克隆抗体通过ELISA与Bt发生反应。在免疫印迹中，单克隆抗体识别出29 kDa和35 kDa的两种蛋白质成分。此外，这些单克隆抗体通过ELISA与Dp提取物发生反应，并且在免疫印迹中检测到相同大小的成分。在竞争性ELISA中，Dp提取物使与Bt抗原的反应性降低了62%，而Bt提取物使针对Dp抗原的反应性最大降低了83%。这些单克隆抗体识别rBlo t1。如抑制试验所示，Blo t1与单克隆抗体识别的蛋白质之间的同源性很高（90%）。
Dp和Bt之间的交叉反应性可能与各自的半胱氨酸蛋白酶等蛋白质的存在有关。所制备的单克隆抗体已被证明有助于鉴定Bt的抗原，并确定Bt与热带和亚热带国家常见螨类群落中其他常见螨类之间可能的交叉反应性，在这些国家中Bt很常见。
