Announcements Quiz Pre lab Lecture More about ELISA Announcements Evaluations WAC partly in class end of Lec 7 Overall subject evaluation available online Mod 3 report No separate methods section needed State anything unique along the way in results section alginate type RNA recovered etc Cross group discussion today or next time ELISA protocol Direct ELISA uses labeled primary antibody Indirect ELISA why use a secondary antibody signal amplification flexibility use 2 with many different 1 efficient but cross reactivity more likely Development process what why how 2 Ab has enzyme AP provide substrate pNPP colorimetric reaction amplification development time is key 3 ELISA Outcomes Outcome Possible Explanations High reading in blank samples cross contamination incomplete washes forgot to block poor systemic binding inhibitor No signal at all old reagents including standards too high Tween wrong 1 or 2 Ab Saturated signal for some samples too concentrated therefore repeat with dilutions MIT OpenCourseWare http ocw mit edu 20 109 Laboratory Fundamentals in Biological Engineering Spring 2010 For information about citing these materials or our Terms of Use visit http ocw mit edu terms
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