How do I avoid impurities from my SEC purification?

More Robert Adamu Shey's questions See All

How do I purify rabbit IgG4?

I want to purify IgG4 antibodies from immunized rabbits. I have tried to look for protocols to no avail. Commercial kits on the other hand exist for humans and a few other animals but I have not...

03 April 2019 10,004 3 View

Any bioinformatics tools to predict immuno-modulatory or immuno-regulatory epitopes in proteins?

Hello everyone, we are characterizing some proteins in the context of vaccine development and we are interested in determining there are any epitopes that may rather lead to immune evasion. Don't...

03 April 2019 2,937 3 View

How do i prepare water saturated phenol:chloroform mixture (5:1)?

I am doing RNA, DNA and protein extraction from the same sample and my protocol recommends this water saturated phenol:chloroform mixture. I am not not very versed with how to go about with the...

03 April 2018 4,314 8 View

What are the best (recommended) tools for 3D protein structure prediction, refinement, and validation; also what is the best tool for MD simulation?

Hello everyone, I am trying to do some hands-on bioinformatics and will be grateful to get your orientation.

31 December 2017 1,350 5 View

How do I interpret protein 3D structure refinement data?

Hello everyone, I did 3D structure prediction (using I-TASSER) for a protein and submitted the best model for structure refinement (with GalaxyRefine). Unfortunately, I don't have a very good...

11 December 2017 1,238 0 View

What is the best buffer to us in antigen-capture ELISAs?

Hello everyone, I am about doing an antigen capture ELISA and a little confused about which buffer can be used in absorbing the capture antibody to the microplates. I am using Nunc MaxiSorb 96...

10 November 2017 2,862 3 View

What is the effects of coating ELISA plates with antigen for longer than recommended durations?

Hello everyone, So I planned this ELISA to do, unfortunately I coated plates and for some reason I am not able to continue with the experiment as planned. So basically, instead of coating...

09 October 2017 9,239 4 View

Autoinduction of pGEX5x3?

Hello everyone, I wish to express a gene cloned in pGEX5x3 through auto-induction and compare with the normal IPTG induction. Any one has experience doing autoinduction with pGEX vectors?

08 September 2017 6,244 0 View

Signal peptides in secretory proteins?

I wish to find out if all secreted proteins have a signal peptide. Also, if yes, are these signal peptides always cut?

07 August 2017 9,895 2 View

Isolation of inclusion bodies?

I am working with a pET30a+ construct and my protein is C-terminal His-tagged. My protein is forming inclusion bodies and I need to get a protocol for isolation of a pure protein from this. An...

07 August 2017 1,962 4 View

Precipitate after adding laemmli buffer to protein solution containing Potassium Acetate?

Hi, I am running a size exclusion chromatography experiment with a buffer containing Potassium Acetate as a salt. I analyse these fractions through SDS-PAGE. After boiling my SEC fractions in...

01 March 2021 2,622 3 View

Are protein samples in SDS compatible with ELISA?

I have used an AllPrep DNA/RNA/Protein Mini Kit QIAGEN kit to extract RNA and protein from my samples. At the end of the protein purification, I resuspend my protein in 5% SDS. Will these samples...

28 February 2021 7,370 3 View

How can I represent SDS-PAGE results for machine learning analysis?

I am trying to classify and analyze the results of an SDS-PAGE based array for bacterial detection using machine learning, but I have trouble finding the best way to represent the results with...

27 February 2021 9,176 3 View

Why my agarose gel staining sometimes looks not homogenic but like gradient (see the figure)?

I am worried about this overexposure of the upper part of the gel in the picture. Is it possible that this is the result of too much concentration of ethidium bromide? Why is there such a big...

25 February 2021 8,140 3 View

How to replicate the well conditions of a crystal hit with 20%(w/v) isopropanol?

I had a crystal hit in a well with 0.1M HEPEs pH 7.5, 10%(w/v) PEG 4000, and 20%(w/v) isopropanol. Why is isopropanol in w/v here if it's just solvent? How do I recreate this well condition?

25 February 2021 9,761 1 View

EMSA troubleshooting. Protein-DNA complex not migrating, cold probe lane with multiple bands. Can anyone help?

I'm running an EMSA to show the DNA binding activity of recombinant proteins versus the Wild type. Previous assays run by my research group using a different test system show the protein-DNA...

24 February 2021 8,325 1 View

Problem with ligating 2.8 kb insert into 8.5 kb plasmid?

I am trying to clone 2 copies of the same mammalian gene into the pSF-CMV-Ub-Puro Ascl (contains HindIII, KpnI and NheI cut sites) plasmid. This is what the final product is supposed to look like...

24 February 2021 6,310 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

Alginate and silk fibroin hydrogels toxic to cells?

Hello, I asm currently working with alginate and silk fibroin hydrogels. I prepared a blend of alginate and aqueous silk fibroin (straight from dialysis) and resorted to ionic crosslinking to...

23 February 2021 8,249 2 View

How to reduce static electricity of powder after lyophilization?

I am working on synthesizing and then purifying peptides using the SPPS system. After purification of the peptide, the peptide powder is obtained using a freeze dryer. When measuring the powder,...

22 February 2021 10,099 5 View

Udaya Jayasundara

SEC separates what you have in your mixture. Probably you may need to treat them prior to inject SEC.

Jasim Al-saadi

You can use Ion Exchange chromatography for the solution resulting from SEC.

Omar Gonzalez-Ortega

If the proteins are not interacting, then you need to improve separation during SEC. Longer column, less volume injected, etc. Do you see only one peak after SEC?