Abstract：
:Protein samples electroblotted onto nitrocellulose membranes and quenched with a mixture of blocking agents produced a strong signal for cystic fibrosis transmembrane-conductance regulator (CFTR), a high-molecular-weight protein, in western blotting. Optimized conditions for CFTR were then extended to medium- and low-molecular-weight proteins (LAMP1 and Rab11a, respectively) to determine the effects of methanol concentration (0-20%) in Towbin's transfer buffer (TTB). Methanol in TTB appears to have little to no effect on CFTR signal. However, for medium-sized (LAMP1) and small (Rab11a) proteins, a lower concentration of methanol (10%) was sufficient to produce a maximal signal. Therefore, methanol, a toxic solvent, can be removed from or reduced in TTB without compromising signal strength. Here, we show modifications that may be useful in detecting and/or improving the signal of low-abundance proteins.
journal_name
Biotechniquesjournal_title
BioTechniquesauthors
Heda GD,Shrestha L,Thapa S,Ghimire S,Raut Ddoi
10.2144/btn-2019-0124subject
Has Abstractpub_date
2020-06-01 00:00:00pages
318-324issue
6eissn
0736-6205issn
1940-9818journal_volume
68pub_type
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