Simultaneous ultrastructural demonstration of multiple peptides in endocrine cells by a novel immunocytochemical method
Simultaneous ultrastructural demonstration of multiple peptides in endocrine cells by a novel immunocytochemical method
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通过新型免疫细胞化学方法同时超微结构演示内分泌细胞中的多种肽
作者：
L. Larsson
目前的免疫细胞化学技术检测所有与组织反应的抗体1。不幸的是，这些抗体中的一些可能与预期抗原以外的抗原反应。通过使用检测所需抗原的不同区域的抗体组来最小化这些问题2，3。然而，由于免疫细胞化学方法现在可以检测到非常低的抗体浓度1，抗血清的纯度至关重要。此外，虽然抗血清可以通过亲和色谱法纯化，但回收高亲合力抗体的困难导致大多数亲和纯化的抗血清富含低亲合力抗体，这些抗体可能在染色过程中脱落1。因此，我们已经开发，并在这里描述，一种新的超微结构后包埋染色技术，基于IgG分子的二价性和使用抗原包被的胶体金颗粒。以前，胶体金标记的抗体已用于包埋后染色4，5。然而，与我们的金标抗原检测（GLAD）技术不同，这些方法不能区分特异性和非特异性抗体。GLAD方法仅检测特异性抗体，不选择高亲合力抗体，在这方面与放射免疫细胞化学方法相似6。然而，GLAD方法与后者的不同之处在于，它可用于超微结构研究，不需要放射自显影，并允许同时检测多种抗原。此外，可以定量与背景相比的比活性。
Current immunocytochemical techniques detect all antibodies that react with tissue1. Unfortunately, some of these antibodies may react with antigens other than those intended. Such problems are minimised by using sets of antibodies detecting different regions of the desired antigen2,3. However, as immunocytochemical methods can now detect very low antibody concentrations1, the purity of antisera is critical. Furthermore, although antisera may be purified by affinity chromatography, difficulties in recovering high-avidity antibodies cause most affinity-purified antisera to be enriched in low-avidity antibodies, which may be dislodged during staining1. We have therefore developed, and describe here, a new ultrastructural post-embedding staining technique, based on the divalency of IgG molecules and using antigen-coated colloidal gold granules. Previously, coUoidal gold-labelled antibodies have been used for post-embedding staining4,5. Unlike our gold-labelled antigen detection (GLAD) technique, however, these methods do not differentiate between specific and nonspecific antibodies. The GLAD method detects only specific antibodies and does not select against high-avidity antibodies, and in this it resembles the radioimmunocytochemical method6. However, the GLAD method differs from the latter in that it is useful for ultrastructural studies, does not require autoradiography and allows simultaneous detection of multiple antigens. Moreover, specific activity compared with background may be quantitated.