Multivalent protein probes for the identification and characterization of cognate cellular ligands for myeloid cell surface receptors.
Cell-cell interactions mediated by cell surface receptor-ligand pairs in the immune system are often of low affinity and transient in nature. To begin to study these weak interactions, it is desirable to devise a generally applicable method for screening for and enriching cells expressing low-affinity ligands for specific cell surface receptors. We describe here an experimental strategy that uses a multivalent form of protein as a probe to identify and characterize cognate ligand(s) of myeloid cell surface receptors. Recombinant fusion proteins containing the receptor protein fragment of interest fused to a truncated Fc domain and a unique biotinylation signal are produced, biotinylated, and coupled to (strep)avidin-coated fluorescent or paramagnetic microspheres. These multivalent microparticle probes are then used to screen or capture cells expressing the cognate cellular ligand(s).
Department of Microbiology and Immunology, Chang Gung University, Tao-Yuan, Taiwan.
Hsi-Hsien
HH
Lin
Gin-Wen
GW
Chang
Yi-Shu
YS
Huang
Cheng-Chih
CC
Hsiao
Martin
M
Stacey
Siamon
S
Gordon
D000818:Animals; D019921:Biotinylation; D016466:CHO Cells; D002130:Calcium Phosphates; D011232:Chemical Precipitation; D002846:Chromatography, Affinity; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA / Q000302:isolation & purification; D005456:Fluorescent Dyes / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D008024:Ligands; D015336:Molecular Probe Techniques; D015335:Molecular Probes / Q000378:metabolism*; D022423:Myeloid Cells / Q000166:cytology* / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D014162:Transfection
D002130:Calcium Phosphates; D005456:Fluorescent Dyes; C029152:G-substrate; D008024:Ligands; D015335:Molecular Probes; D009419:Nerve Tissue Proteins; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; D004247:DNA; C020243:calcium phosphate