Cell Signaling Technologies
LC3B XP® Rabbit mAb
3868
(D11)
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We wanted to be able to quantify the ratio of LC3B-I and LC3B-II. The sizes are 16 and 14 kDa respectively and are quite hard to resolve these two bands. With some modifications to our normal blotting protocol, we were able to achieve these results.
Western Blot
BMDMs
1:1000 overnight at 4 degrees C
5% BSA
1:3000 anti-rabbit HRP linked for 1 hour at room temp
None
SuperSignal™ West Femto Maximum Sensitivity Substrate
We were able to resolve the 2 bands after optimizing our protocol.
We used 8-16% gels and ran until 1the 10 kDa marker was nearly off the gel (the loading dye had run off). We used a rapid transfer optimized for low MW proteins.
Good detection, clean blot
Can be hard to optimize
With some modifications to our normal blotting protocol, we were able to identify and resolve the two types of LC3B in order to quantify the ratio.