I'm currently experiencing a lot of problems using the BIACORE T100 in my lab. I'm working with an RNA binding protein which also binds to ssDNA. My current setup is:
SA chip immobilised with biotin-DNA
Buffer: 10mM Tris-HCl at pH 7.0, 150mM NaCl (from DEPC treated H2O) [0.025% P20, 1mM DTT added fresh] Buffer is degassed and filtered before use.
My first time with BIACORE, a few injections had small spikes on the sensorgrams and baseline did not return to starting position.
Before my second run with BIACORE, while trying to reach baseline, little spikes showed up at about every 15 seconds - not sure why this is happening.
So we decided to get the BIACORE machine serviced about a month later. It prompted me for a desorb and sanitize so I ran that.
This time, when I ran my experiments, blank injections were showing negative response units at about -100RU ... and the shape of the curve is such that there's a second dip in the 2nd half of the inject. We thought that it may be because the chip was sitting idle for a month, so we decided to use a new chip and immoblise it with DNA again. We ran the experiment again and the blanks came out the same. Previously, my protein had nM affinity. nM injections of protein give a negative RU, and injections of concentrations > 10uM give a positive RU.
Has anyone experienced problems like this or have any suggestions as to what I may be doing wrong?
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