By Richard Horuk (Eds.)
Chemokines are significant mediators of immune cells and are enthusiastic about quite a lot of proinflammatory human ailments, together with rheumatoid arthritis, a number of sclerosis, and organ transplant rejection. It has lately been found that their receptors are fascinated about HIV an infection. The characterization of those molecules and their receptors is therefore of fundamental value in knowing a few human illnesses and infections. This quantity and its significant other quantity 287 Chemokines offer accomplished experimental protocols utilized in this box of analysis.
* Chemokine receptors
* Chemokines in disease
* sign transduction of chemokines
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Extra resources for Chemokine Receptors
Stably transfected 293 cells expressing IL-8RA (293-IL-8RA) or IL-8RB (293-27) are generated using clones described previously, n'12 Briefly, 293 cells (a human fetal kidney cell line) are cotransfected with 11 W. E. Holmes, J. -J. Kuang, G. C. Rice, and W. I. Wood, Science 253, 1278 (1991). 12j. Lee, R. Horuk, G. C. Rice, G. L. Bennett, T. Camerato, and W. I. Wood, J. Biol. Chem. 267, 16283 (1992). p. p. Sera adsorbed with 293 Tested on 293-71 by FACS + I F os I Inhibition of*lL-8 binding to 293-71 ~1 5+ / 86 / Inhibition of *IL-8 binding to 293-71 FACS 293-71 and 293 4, J Non-cross-reactivity 293-71 (a) vs 293-27 (b) 2+/2 F ~ .
N H ~ - - ~ 190~ ~280 03 | 154 ~--COOH 349 Fia. 1. Model of the secondary structure of human interleukin-8 receptor A. Dots represent potential glycosylation sites. block the ligand binding to a particular receptor would be extremely useful for understanding the biology of each ligand and the distribution and regulation of its receptor expression. Although much has been learned since the first report of monoclonal antibodies by Kohler and Milstein,3 the generation of MAbs to chemokine receptors has remained a challenge.
288, 182 (1997). 3. The fixative is removed after 1 hr, and the cells are carefully washed twice in PBS and then stored in PBS at 4 ° in an airtight container until the time of immunostaining (never store for longer than 2 to 3 weeks unless PBS is replaced because evaporation will alter osmolarity). 005% Triton X-100 (v/v) and 10% (v/v) serum from either goat or donkey (depending on the species of the secondary antibody). The blocking solution is removed, and the primary antibodies (diluted in the same buffer used for blocking) are added without a rinse step.
Chemokine Receptors by Richard Horuk (Eds.)