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).
Lin
Hsi-Hsien
HH
Department of Microbiology and Immunology, Chang Gung University, Tao-Yuan, Taiwan.
Chang
Gin-Wen
GW
Huang
Yi-Shu
YS
Hsiao
Cheng-Chih
CC
Stacey
Martin
M
Gordon
Siamon
S
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