Methods in Membrane Biology: Volume 9 - download pdf or read online

By S. de Petris (auth.), Edward D. Korn (eds.)

Many of the equipment now generally use in membrane biology, and never already mentioned in passable aspect in other places, were lined within the 8 formerly released volumes of this sequence. a lot of this 9th quantity is occupied by means of one authoritative bankruptcy, an strangely thorough and demanding evaluation of a comparatively new and hugely really expert know-how that has received quick recognition: immunofluorescence and immunoelectron microscopy. those are strong experimental instruments appropriate in fields a lot broader than membrane study and using equipment drawn from extensively varied disciplines reminiscent of natural chemistry, protein chemistry, immunology, and fluorescence and electron microscopy. The temptation to take advantage of those great­ ficially, and deceptively, basic yet essentially advanced tools un­ significantly is sort of overwhelming. The bankruptcy through de Petris, a pioneer within the box, is as worthy because it is rigorous, and it's going to lengthy be the normal during this region of analysis. the second one bankruptcy during this quantity is a extra really expert assessment via Matus of the systems for the education and characterization of the hugely differentiated junctional areas of mind plasma membranes. those equipment are relevant to the speedily starting to be box of neurobiochemistry­ membrane biochemistry at possibly its so much intricate.

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Fig. 8. Examples of direct lectin receptor labeling with ferritin-conjugated concanavalin A. Stained sections (as in Fig. 7B). Continuous ferritin layer in a cap of concanavalin A receptors cross-linked by the conjugate on a mouse thymocyte. The distribution of the label is in general more regular than that of antibody conjugates, and the ferritin molecules are often at a nearly constant distance from the membrane (10-12 nm) (cf. also Fig. 9). The ferritin cores are clearly visible also on membrane areas cut tangentially to the plane of the section.

150) (L. Wofsy, C. Henry, and S. Cammisuli, personal communication). This is a simplified version of the hapten-bridge methods described in the following section. The method, which requires the extra step of the coupling of the hapten to the antibody, does not present any advantage over the simple two-layer 34 S. de Petris method, which employs unmodified antisurface antibodies, if only one antigen has to be labeled. The method, however, may be useful for labeling simultaneously two different antigens using surface antibodies coupled to different haptens (see below).

Purified antibodies are required in the direct method only when the marker is an enzyme (Avrameas, 1970; see, however, Nakane and Kawaoi, 1974). The direct method also has the advantage that the number of bound marker molecules is directly proportional, and often nearly equal, to the number of the bound antisurface antibody molecules. Using ferritin as a marker, the method (together with the two-layer hybrid antibody method) also has the highest resolution for the localization of surface antigens.

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