Immunoblotting

Immunologic method used for detecting or quantifying immunoreactive substances. The substance is identified by first immobilizing it by blotting... | Contact experts in Immunoblotting to get answers

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Questions related to Immunoblotting

Can someone offer an explanation for two methods of Immunoblotting phosphorylated proteins?

I'm looking to detect pFAK Tyr397. I've found two methods, the first is to run whole lysates on a gel, and blot for a pFAK antibody (ie pFAK-397) and the other is to IP the FAK out of the lysate,...

11 November 2015 8,876 5 View

Does the increase in cardiolipin content correlate with mitochondrial fission process?

What i mean to ask is whether an increase in fluorescence of nonylacridine orange staining along with an increase in mitotracker red fluorescence may be attributed as Mitochondrial fission? Of...

10 October 2015 3,010 3 View

Why is my Western blot with human striatum for ceruloplasmin not working?

I am doing western blots on human striatum for ceruloplasmin but cannot get any bands. I know that the concentration of ceruloplasmin in the brain is quite low. So I've tried using a higher...

09 September 2015 9,615 4 View

What is a good Schlafen 9 antibody for immunoblots?

I am trying to find some good antibody for schlafen 9 to be used for western blots.Please suggest me some good schlafen 9 antibody. Thanks and Regards

09 September 2015 1,035 1 View

When doing an immunoblot, what would cause the dye front, standards, and protein to all run together when they reach the bottom third of the gel?

I have had it happen to me three separate times now that when running a 12% SDS-PAGE gel, the dye front stalls/slows two-thirds through the gel and the ladders and my protein eventually catch up...

08 August 2015 1,773 3 View

Does anyone know (or has) a good anti-mouse-IL-10R for western and IP?

We are interested in assessing the kinetic of binding of STAT3 to the IL-10R in mouse T lymphocytes, but we are having a hard time finding a reliable anti-mouse-IL-10R antibody (that possibly...

07 July 2015 815 3 View

Should I double-label alpha-SMA with E-cadherin or B-catenin when looking at an epithelial-mesenchymal transition (EMT) response?

I'm planning to do both immunofluorescence and Western Blot analysis and am wondering which membrane marker would be the best to use. It is well-established that B-catenin gets translocated to the...

07 July 2015 1,815 4 View

Why my ubiquitination detection given only one band?

I have been tested ubiquitination of one enzyme. In our lab ,we have one cell line overexpressed this gene( with his and myc tag). So I used myc to IP this enzyme and then use WB to test if it...

07 July 2015 5,602 15 View

Any suggestions to immunoblot S307 IRS in C2C12 cell lines- both basal and insulin stimulated?

I used Ser 307 IRS antibody at a concentration of 1:1000 and it was purchased from CST as IRS antibody sampler kit.

07 July 2015 9,569 4 View

Can we use horizontal gel electrophoresis for western blotting?

Western blotting

06 June 2015 9,051 3 View

How long are brain samples stored at -80 good for (pre-lysing), for western blot analysis?

I have samples from a year ago and wondered if they would still be able to be processed and used.

06 June 2015 3,958 3 View

Does anyone know about washing western blot membrane w/ TBST overnight?

so the primary antibody we're using is particularly dirty, and even with 5+ five minute washes in TBST, we still get a good amount of background. I know its not the blocking or secondary. I ...

06 June 2015 5,570 5 View

How can I calculate the volume of sample to be loaded with desired concentration in wells after DC Assay?

Hi Everyone, Can anyone help me in calculating the lysate volume of required concentration to be loaded in wells after DC Assay for western blot analysis?

06 June 2015 4,276 1 View

How can I localize a protein, detected by 2D immunoblotting, on the silver staining gel?

I have been struggling in trying to localize the protein of interest i detect by 2D immunoblotting on duplicated silver staining.i tried to duplicate the 2 D gel and use one for 2D Immunoblotting...

06 June 2015 8,117 4 View

I would like to know what is prefer BSA or nonfat dry milk in western blot?

I would like to know which is prefer BSA or nonfat dry milk to block nitrocellulose membrane.

05 May 2015 4,420 7 View

Is it possible to conclude about mitochondrial fission basing on phosphorylation of DRP1 and TEM images?

I would like to assess whether my experimental protocol affects mitofusion/mitofission rate in tissues in vivo. I prepared immunoblots with antibodies against phosphoDRP1. I also plan to prepare...

05 May 2015 9,991 11 View

Which is the best GluR1 antibody to detect GluR1 in IF and Western Blot?

I would like to detect a quantitative change in Western Blot and IF which I have detected with mass spectrometry.

05 May 2015 9,097 1 View

Can probing multiple primary antibodies together in one membrane result in low or no band signal in WB?

Hello, I incubate two type of antibodies A and B together in the same membrane. Both are same species monoclonal, but bind to different protein molecular weight. It usually give me the result in...

05 May 2015 1,507 4 View

Can anybody tell me what are the other bands in my immunoblotting with 6E10 antibody (covance) . See the pic for details

Alzheimer's disease

05 May 2015 5,301 10 View

Why is the band distorted in this western blotting?

I have tried alot and don't know the reason why these bands are distorted? Can any one suggest me? the pic is attached below.

05 May 2015 1,167 15 View

I would like to know what is prefer BSA or nonfat dry milk in western blot?

I would like to know which is prefer BSA or nonfat dry milk to block nitrocellulose membrane.

05 May 2015 9,479 6 View

Which is the best GluR1 antibody to detect GluR1 in IF and Western Blot?

I would like to detect a quantitative change in Western Blot and IF which I have detected with mass spectrometry.

03 May 2015 450 1 View

Has anyone analyzed porcine Cytochrome C via Western Blot?

We are looking for a primary antibody against cytochrome c known to work in pigs. Thank you!

14 April 2015 3,570 3 View

How can I make immunoblot bands straight and not curved?

Immunoblot bands of protein size below 20KDa (19, 15, 12) becomes curved, I use PC9 cell. However bands for larger protein size has no problem. I use 12% and 15% SDS Gel and gel running at 100...

04 April 2015 5,132 1 View

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