Non-specific bands in PCR?

Usually I am doing PCR with 0.5 ul of Taq polymerase (abm) per reaction. Since I wanted to increase the sensitivity of target amplification, I tried PCR with 1.0 ul of Taq. It gave some bands in...

02 March 2018 9,088 6 View

Mass spectroscopic analysis for protein identification?

I am doing immunoprecipitation for concentrating by target proteins. I usually elute my proteins in SDS loading buffer containing Tris, glycerol, SDS, beta mercaptoethanol and bromophenol blue....

11 December 2017 1,141 2 View

Human Anti-Leishmania IgG ELISA Kits...

I have found two commercial ELISA kits for detection of human leishmaniasis. 1. Human Anti-Leishmania IgG ELISA Kit (ab108749) by Abcam 2. Leishmania IgG ELISA Kit (KA3295) by Abnova Both are...

03 April 2017 3,589 1 View

Why did the protein sample and marker not migrate properly in the separating gel (SDS-PAGE)?

I am facing the a problem in my SDS page gel. protein marker and samples not get well separated. All samples were stop separating at the middle of separating gel (protein marker also). Dye front...

09 October 2016 3,280 12 View

Physical characterization of crude parasite Ag???

Is there a simple method to physically characterize the crude Ag extracted from a parasite to check whether they are proteins, polysaccharides or phospholipids like compounds? I am using crude Ag...

07 August 2016 3,925 3 View

Analysis of proteins using westernblotting?

Does 3D structure of a protein affect for migrating distance of a protein?

06 July 2016 6,207 0 View

What is the best condition for blocking western blot membrane?

Can I keep my blocking buffer (skimmed milk or BSA) at room temperature for overnight? Is that affect for my protein?

05 June 2016 3,857 5 View

Commercial protein sequencing services?

I want to sequence some unknown protein excised from 1D-SDS-gel. Have anybody experienced with protein sequencing? Are there any good commercial protein sequencing services?

04 May 2016 4,446 1 View

Fuzzy bands on SDS-PAGE?

What are the reasons for getting fuzzy bands on SDS-polyacrylamide gel?

03 April 2016 463 9 View

TMB SUBSTRATE FOR WESTERN BLOT???

How can we convert TMB of ELISA for western blot by converting soluble TMB product of ELISA into insoluble with addition of 0.05% sodium nitroprusside? Can I use ELISA TMB substrate for Western blot?

02 March 2016 8,565 0 View

Why does blocking with my immunogenic peptide cause nuclear fluorescence in ICC?

Hi all, I am validating a polyclonal antibody raised against human ANT4, through WB and ICC. In most of my controls, the antibody shows very little off-target binding, but when I pre-incubate the...

03 March 2021 5,999 3 View

RIA. How to measure?

When you use RIA, with a control the unknown sample with antibodies, you bare in mind the binding sites of the antibodies. However you still need to measure labelled antigen (radioactive) and not...

28 February 2021 2,133 3 View

Will a 10xHis-tag prevent anti-MBP from binding to MBP?

I have a protein with a 10xHis-MBP tag on the N-terminus. I'm fairly certain I am expressing my protein, but I'm not getting a signal on my western blot. Is there a possibility that the 10xHis tag...

25 February 2021 8,404 3 View

What's the concentration recommend for recombinant protein use in western blotting?

I'm currently working having problem with the concentration for recombinant protein use in western blot, the one been used is Recombinant Human CD19 Fc Chimera Protein-rndsystem which does not...

24 February 2021 8,514 3 View

Standard curve shifting over time. How do I stabilise the ELISA?

24 February 2021 1,365 3 View

What is the best way to manage Western blots?

Increasingly, more articles are being retracted because of duplicate or otherwise dubious Western blots. Consequently, journals are asking for original Western blots and authentication of...

23 February 2021 8,609 3 View

How to manage western blot for proteomics data validation?

Hi Everyone, I am a bigginer in western blotting and I am trying to use it for validation of my proteomics data. I have: two regulated proteins with molecular mass of 40 and 73 kDa one control...

23 February 2021 6,671 4 View

Diltuions for a Western blot?

I need help using the equation C1V1=C2V2 figuring out how much antibody to add to use for a western blot. The stock was origianlly 3.45 ng/ul but diluted 1:10 so now its 0.345 ng/ul. I need to run...

22 February 2021 9,901 3 View

Can anyone help me how to get the similar bands in Actin?

I run western blot on HCT-116 cell lines, I got actin band like in the attached picture. The band size is not same. Can anyone help me explain this situation and how to overcome? Thank you your...

22 February 2021 6,937 3 View

I see 2 random bands in my ERK/pERK blot? Has anyone else seen this?

I ran a western blot using 12% gel and probing for ERK/pERK (cell signaling antibodies). The green is total ERK, the Red is supposed to be phospho-ERK. I see 2 bands below the total ERK that...

22 February 2021 1,777 3 View