BIOL 620 121 1st Edition Lecture 5Outline of Current Lecture Immunology Lecture 5- Avidity and classes of immunoglobulinsCurrent Lecture Avidity is the strength of binding between polyvalent Ab and polyvalent Ag.Avidity= Ka=affinity constant.If there is higher Ag binding site, Ka is lower, but the avidity is high. In other words, it is the number of binding sites that count.Equilibrium dialysis: Radio- labeled haptens are used to measure radiation. Measurements of Ab binding to hapten, amount of free haptens, and free Ab are taken into consideration.Side A: radiation from free hapten and bound hapten.Side B: radiation from free hapten only.Ratio: total bound hapten/ total Ab. Or Difference in radiation between Side A and Side B/ initial added. Many experiments are done where the Ab concentration is kept constant and the amount of hapten is varied. In another set of experiments, Ab is kept constant and hapten concentration is varied. The graph is plotted where x- intercept gives the valence.The graph is a curve due to the polyclonal nature of Ab as it recognizes different epitopes. Slope of the tangent gives Kav. If the slope is steep, it denotes higher affinity. Higher avidity refers to binding site and its functions. Even a single response to Ag can produce more Ab and more Ab are better in affinity. This is referred to as affinity maturation.Experiment: Ova albumin immunized into rabbit - take serum -- gel electrophoresisThe immune serum has no gamma globulin band. In other words, the experiment provesthat Ab are present in gamma –globulin fractions and are called as immunoglobulins, Ig.Antibody classes: Serum levelIg G- 150 K Da 13 mg/mlIg A 3 mg/ mlIg M- 900 K Da 0.3 mg/mlIg D 0.03 mg/mlIg E 3 X 10^-4
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