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Trouble with Your Westerns?

Click for some time (and money saving tips)!

Improving your Western Blots

Seven tips to improving your Western Blots:

·         1. Uniformly distributed high background?
o   You may have insufficiently washed.  Increase the number of washes and buffer volume. 
§  Make washing easier with  Blot Boxes.
o   Antibody concentrations too high
§  Make sure to optimize your antibody concentrations by running strips at varying concentrations to determine best concentration for your samples.

·         2. Weak or no signal?
o   Your protein may have not transferred well. 
§  Check your transfer buffer make-up and blotting procedure.  Stain gel to visualize proteins remaining in gel. 

·         3. The Ladder Lane is black
o   The antibody may be reacting with your MW (ladder) marker.
§  Add an empty lane between the MW marker and sample lane.

·         4. The band of interest is too low/too high on the blot.
o   Separation is not efficient. 
§  Change the gel percentage: A higher percentage for small protein, a lower percentage for larger proteins. Or try a  gradient gel.

·         5. Ripped resolving gel when moving to the cassette for transferring?
o   Try Run Blue™ pre-cast gels from MIDSCI.  They are stronger and resist tearing! No more piecing together torn gels!

·         6. Sample bleeding over to another well or difficulty loading?
o   Try the deeper wells from Expedeon!  Easily hold 35ul of sample without bleeding over to another well!
o   Try gel-loading tips.  Available in flat or round, they make gel loading easier!

·         6. Worried about pre-cast gels expiring or losing resolution before you can use them?

o   Expedeon Run Blue™ gels have a 2 year shelf life with no loss in resolution.  Use them confidently from purchase date to 2 years later!

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