Competition BIAcore for measuring true affinities: large differences from values determined from binding kinetics. Export

@article{citeulike:472096, abstract = {In attempting to use the BIAcore instrument for the determination of binding constants of several haptens or peptides to different antibodies by measuring on- and off-rates, we found that neither the absolute nor the relative values of the binding constants corresponded to the measurements in solution. Even at the lowest coupling densities useful for measurements, rebinding and bivalency effects offset the measurements by a factor of up to 500. We caution therefore about using on- and off-rates for the determination of absolute or even relative binding constants without controlling for rebinding and avidity effects. Instead, we show that binding constants in solution can be reproduced well by using on-rate determinations of antibody preincubated with antigen, and we derive the conditions under which such an approach is valid.}, address = {Biochemisches Institut der Universit\"{a}t Z\"{u}rich, Switzerland.}, author = {Nieba, L. and Krebber, A. and Pl\"{u}ckthun, A.}, citeulike-article-id = {472096}, citeulike-linkout-0 = {http://view.ncbi.nlm.nih.gov/pubmed/8714593}, citeulike-linkout-1 = {http://www.hubmed.org/display.cgi?uids=8714593}, citeulike-linkout-2 = {http://dx.doi.org/10.1006/abio.1996.0067}, day = {15}, doi = {10.1006/abio.1996.0067}, issn = {0003-2697}, journal = {Anal Biochem}, keywords = {affinity, analysis, antibody, spr, statistic}, month = {February}, number = {2}, pages = {155--165}, posted-at = {2008-12-02 09:34:07}, priority = {2}, title = {Competition BIAcore for measuring true affinities: large differences from values determined from binding kinetics.}, url = {http://dx.doi.org/10.1006/abio.1996.0067}, volume = {234}, year = {1996} }

TY - JOUR ID - citeulike:472096 L3 - citeulike-article-id:472096 N2 - In attempting to use the BIAcore instrument for the determination of binding constants of several haptens or peptides to different antibodies by measuring on- and off-rates, we found that neither the absolute nor the relative values of the binding constants corresponded to the measurements in solution. Even at the lowest coupling densities useful for measurements, rebinding and bivalency effects offset the measurements by a factor of up to 500. We caution therefore about using on- and off-rates for the determination of absolute or even relative binding constants without controlling for rebinding and avidity effects. Instead, we show that binding constants in solution can be reproduced well by using on-rate determinations of antibody preincubated with antigen, and we derive the conditions under which such an approach is valid. IS - 2 JF - Anal Biochem SN - 0003-2697 AD - Biochemisches Institut der Universität Zürich, Switzerland. EP - 165 TI - Competition BIAcore for measuring true affinities: large differences from values determined from binding kinetics. VL - 234 SP - 155 KW - affinity KW - analysis KW - antibody KW - spr KW - statistic AU - Nieba, L AU - Krebber, A AU - Plückthun, A PY - 1996/02/15/ UR - http://dx.doi.org/10.1006/abio.1996.0067 ER -