http://biowww.net/ Tue, 15 May 2012 05:22:35 +0100 FeedCreator 1.7.2 http://biowww.net/detail-262.html Preparation of cell lysates from E.Coli bacteria for western blotting: 1. Spin down 1 ML overnight culture 2. add 100-200 ul BugBuster protein extraction reagent (Novagen cat. No. 70584) 3. Sonicate 1 min 4. add 2x sample buffer to same lysis volume(or a ... biowww http://biowww.net/detail-1377.html A technique note on multiplex western blot analysis using Qdot fluorescent technology. Nature Methods 2, 79 - 81 (2005) Abstract: Western or immunoblotting analysis of protein expression in cells and tissues has been the major analytical tool for as ... biowww http://biowww.net/detail-1350.html Western blot stripping protocol: Buffer needed: stripping buffer: 2% SDS 100 mM beta-mercaptoethanol 50 mM Tris, pH 6.8 Procedure: 1. Heat stripping buffer to 50c in water bath. 2. Incubate blot with stripping buffer at 50c for 15-30 min with gen ... biowww http://biowww.net/detail-1351.html Semi-dry western blot transfer procedure: This protocol describes the procedure for transfering protein from polyacrylamide gel onto nylone membrane using semi-dry transfer method. 1. Cut 6-10 sheets of whatman 3MM the same size as the mini PAGE gel. ... biowww http://biowww.net/detail-395.html A typical western blot procedure includes following steps: 1. Protein lysate preparation from cells and tissues. 2. Protein assay for determination of protein concentration in lysate. 3. Denaturing and reduction of protein complexes before loading onto ge ... biowww http://biowww.net/detail-380.html A newsgroup post on large protein transfer efficiency for western blot. I have been having a lot of problems with efficiency of transfer from gel (10% acrylamide/bis) to PVDF membrane (Immobilon P). When I stain the gel after transfer, there is a lot o ... biowww http://biowww.net/detail-49.html Company kits end up costing $3-5 per western blot which is 90% of the cost of the whole procedure. People must have some good homemade HRP (and AlkPhos) Chemiluminescent Detection protocols or references? ... ... Click follow ... biowww http://biowww.net/detail-77.html Does anybody know if it makes a difference for certain antibodys (for protein-detection on a western-blot) to use PBS instead of TBS as basic for all the incubation- and washing-buffers? The reason why I´m asking is the following one: ... ... biowww http://biowww.net/detail-104.html I've had a couple of weird western blots lately, we use a standard SDS PAGE tris glycine minigel (BIORAD), and transfer using the minigel apparatus at 100v for 90 minutes with a cooling block (ice in a container) present. The transfer buffer is ... biowww http://biowww.net/detail-133.html I have been having a lot of problems with efficiency of transfer from gel (10% acrylamide/bis) to PVDF membrane (Immobilon P). When I stain the gel after transfer, there is a lot of protein left on there. I know that I am not exceeding the binding cap ... biowww http://biowww.net/detail-401.html From Roche Applied Biosciences (PDF file), ... biowww http://biowww.net/detail-469.html From R&D systems. It gives troubleshooting guide on several common problems on western blot: low protein transfer, high background, low protein binding, multiple bands staining etc. ... biowww http://biowww.net/detail-653.html A FAQ from Synaptic System on problem of many bands on a western blot. Beside solutions provided above, some other considerations may be helpful for eliminate multiple bands in western blot. Some solutions for high background of western blot (multiple ba ... biowww http://biowww.net/detail-1137.html Western blot troubleshooting tips - protein transfer problem: poor transfer efficiency, air bubbles trapped when assembling the transfer sandwitch (use a tube or roller to get ride of any trapped air bubbles), voltage and current is not optimized for ... biowww