Western Blot

Identification of proteins or peptides that have been electrophoretically separated by blot transferring from the electrophoresis gel to strips of... | Contact experts in Western Blot to get answers

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Questions related to Western Blot

Why does fluorescent WB detection fail while chemiluminescence is fine?

I have a simple western blot of CD31 in HUVEC lysate. With an anti-mouse-HRP secondary Ab I get a good signal/band. With fluorescent secondary antibody there is no signal. I have tried multiple...

06 August 2019 6,186 3 View

Why am I getting lots of background dots on nitrocellulose membranes?

Edit: I have now managed to attached examples. These are both the same membrane, just different channels. I only see the dots when I use rabbit-800 secondary ab, and am now thinking it could be...

01 August 2019 3,156 12 View

What are the white bands on western blot ?

Out of curiosity ! I was trying to do western blot for Erythropoietin, as you can see the procedure was unsuccesful but there are some unexpected white bands on the blots. Can anyone tell me what...

30 July 2019 5,800 13 View

I performed P-AKT western blot experiment. Even in 0 MOI, I got band. Is there any suggestion?

In any other phosphorylated proteins, in 0 MOI, I did not get band, but I got band in P-TAK.

23 July 2019 1,988 5 View

Why I have very strong phospho protein signal in western blot?

I am working with phospho protein in western blot and have problems about the phospho protein signal. It is common that researchers can not detect phospho-protein signal in the membrane, as...

18 July 2019 6,149 3 View

Is a non-specific protein getting pulldown in my Co-IP experiment? If it is then how do I avoid this protein?

I am trying to pulldown a protein of 300 kDa using protein G agarose beads. I used two negative controls, one pulldown with just beads (without primary antibody) and another pulldown with another...

14 July 2019 3,373 8 View

Please could someone help me with the interpretation of a Western Blot?

See attached photo of the Western Blot, I am new to the field and am struggling with interpreting the Blot, any advice would be appreciated.

12 July 2019 3,248 3 View

Do you have any suggestions or protocol?

I am interested in studying a membrane protein in insect larvae (prohibitin). I'm having trouble visualizing it in western blot. When the protein extract is stored in freezing, when it thaws,...

10 July 2019 4,949 3 View

Apart from western blot, what are the other tests that can be used to determine expression of autophagy-related proteins ?

I would like to know if there are other options that requires less cost.

10 July 2019 7,112 2 View

How can you stain unstained ladders PRIOR running gel electrophoresis for western blots?

Good day, I have seen that our laboratory has a lot of stock of Precision plus unstained standards and very few of Precision plus stained standards. Since I need to run a lot of western blots in...

10 July 2019 2,696 2 View

Western blot: with high background, multiple nonspecific white bands along with marker?

I am doing western blot for my samples and every-time getting high background with multiple nonspecific white bands, even marker gets stained as white bands. I have tried all different parameters...

08 July 2019 5,176 6 View

Do you have any suggestions or protocol?

I am interested in studying a membrane protein in insect larvae (prohibitin). I'm having trouble visualizing it in western blot. When the protein extract is stored in freezing, when it thaws,...

07 July 2019 4,210 4 View

Apart from western blot, what are the other tests that can be used to determine expression of autophagy-related proteins ?

I would like to know if there are other options that requires less cost.

07 July 2019 8,413 3 View

Is Click-IT AHA compatible with ELISA detection?

I am trying to measure protein synthesis rate of a particular secreted protein that is very hard to see in western blot. However we have a good stablished method for ELISA detection. My question...

07 July 2019 8,473 3 View

Which running buffer is more compatible to detect TSG101 for exosomes?

I am trying to standardize exosome isolation protocol and want to run a western blot to detect TSG101. Currently, our company does not have a Tris-glycine based system and have been using MOPS...

07 July 2019 1,086 2 View

Anti-human DP1 antibody?

Can anyone recommend a good anti-human DP1 (type-1 prostanoid receptor) antibody for either flow cytometry use? I appreciate there might be antibodies out there not validated for flow cytometry so...

07 July 2019 9,460 5 View

Why spotty bands for western blot?

Hello everyone, I practiced protein extraction for my non-senescent and senescent HDF samples and performed western blot. My results show spotty bands for caveolin1. My senior says that I got some...

07 July 2019 1,098 14 View

What is the best Antibody for detecting Human Growth Hormone with western blotting technique?

Hello, I am looking for a suitable antibody for hGH. I wanna to confirm my result with a confirmatory test such as Western Blot. Thank you.

07 July 2019 8,098 3 View

How to lysis an expressing a membrane protein using a plasmid?

I have created a plasmid for my protein of interested tagged with Flag tag. The sequence looks fine but for the life of me I cant get this protein to express itself and see it using western blot (...

07 July 2019 4,837 16 View

Western blot: with high background, multiple nonspecific white bands along with marker?

I am doing western blot for my samples and every-time getting high background with multiple nonspecific white bands, even marker gets stained as white bands. I have tried all different parameters...

07 July 2019 2,969 7 View

How can you stain unstained ladders PRIOR running gel electrophoresis for western blots?

Good day, I have seen that our laboratory has a lot of stock of Precision plus unstained standards and very few of Precision plus stained standards. Since I need to run a lot of western blots in...

07 July 2019 7,373 2 View

Can you load different amount of samples on western blot?

Usually I load about 20ug on the each well, but I was thinking if it would be a good idea to load protein in such way that well1: 10 ug well2: 20 ug well3: 30 ug .... and so on. Would the band...

07 July 2019 6,978 5 View

I performed P-AKT western blot experiment. Even in 0 MOI, I got band. Is there any suggestion?

In any other phosphorylated proteins, in 0 MOI, I did not get band, but I got band in P-TAK.

07 July 2019 3,105 6 View

How we can conclude that the drug can affect to the apoptosis of the cell through intrinsic pathway?

I have 2 cell lines ( cancer cell lines) and a compound. First, I perform the MTT assay to identify the IC50 of 2 cell lines (~5 micro molar). Then the flow cytometry was perform to see that this...

07 July 2019 8,390 4 View

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