Immunoprecipitation: beads without antibody binding still give a signal?

Hi all, As a negative control, I've used a set-up in which: 1) beads without antibody binding were incubated with the sample and then put on WB. I don't see any heavy chain (which is logic),...

01 February 2020 7,255 2 View

Slower migration on SDS-page in complex sample?

Hi all, Is it possible that the migration of my protein is delayed due to the complexity of the sample? When I run a western blot with 1) a protein extract from locust tissue and 2) hemolymph...

05 June 2019 2,727 11 View

Secondary antibody gives same signal as primary?

Hi, After immunoprecipitation and elution, I check the eluate with western blot. If I incubate the blot ONLY with secondary antibody, it gives the same signal as incubatin with primary &...

31 December 2018 1,079 5 View

Western blot on immunoprecipitation eluate detects related protein?

Hello, So, first I perform immunoprecipitation. The eluate is then subjected to immunoblot analysis using an antibody against a related protein (equal size) and gives a band. How sure am I the...

10 November 2018 4,435 8 View

Allways same (wrong) sequence after sanger sequencing?

Hi, I've cloned and sequenced my PCR product but, besides a good run, I allways find the same (wrong) sequence in other runs. Running a BLAST gave no result. What could this be?? Thanks!

01 February 2018 3,433 0 View

Only cds on NCBI?

Hi, When you look up an mRNA seq of a certain gene, it says "complete cds" on NCBI. This means that only the coding sequence is given, right? When you compare this mRNA seq to a NCBI TSA...

08 September 2017 7,470 2 View

How to separate a complex fluid in a vesicular and non vesicular part

Hello, Is it possible to separate a complex fluid in a vesicular part (containing vesicles / exosomes) and a non - vesicular part, containing other proteins? Many thanks!

05 June 2017 2,792 4 View

What is the function of XBP buffer in exosome isolation?

04 May 2017 1,036 2 View

Synthesis of RNA mimics/antagomirs?

Hello, I was wondering how RNA mimics and antagomirs are being synthesized. I don't need the details (volumes etc), but just a general overview of enzymes being used etc? Thanks! Cheers

09 October 2016 2,398 1 View

Different elution methods for immunoprecipitation result in different bands on western blot?

01 January 1970 5,929 6 View

Gel electrophoresis result for total RNA samples, 1.5 Agarose gel was used, How to improve the integrity of RNA?

Gel electrophoresis, RNA degradation, RNA extraction from fresh tissue

02 March 2021 5,433 5 View

Problem with shRNA transfection. Why are only two plates getting contaminated instead of all?

I transfected my LNCaP-WT cells with 3 shRNA plus their NTC two weeks ago and split two puromycin selected cell plates on Friday last week(Feb 26). I checked for GFP in the cells, and they all...

28 February 2021 4,949 3 View

Any specific pipeline or parameter to analyse total transcriptomics study of brain tissue samples?

I have two groups of brain samples, control and treated for example. It was total RNA nova seq sequencing. I tried all the available pipeline like: star+rsem+deseq2, Hista+stringtie+cuffdiff,...

27 February 2021 356 6 View

Why is the efficiency of viral mRNA electroporation in PBMC low?

Hello, I have been struggling with the electroporation of PBMC with HIV-I mRNA. Although the quality of the RNA generated is good, and the handling of the cells in meticulous, the frequency of...

27 February 2021 9,233 3 View

What is the best concentration of cDNA to be used for qPCR?

I have synthesised cDNA from 500ng/ul of RNA. The concentration of the cDNA Is about 5000ng/ul. Should I dilute my cDNA or just use it as it is for my qPCR? I usually use 2ul of cDNA for a 20ul...

24 February 2021 8,618 3 View

Recommendations on homogenizers for RNA isolation?

I will be performing RNA isolation with frozen lung biopsy samples and am in the market for a tissue homogenizer. I am wanting a homogenizer that will fit in a 1.5ml tube. I was looking at the...

24 February 2021 3,623 3 View

RNA-seq DEGs downstream analysis and reporting in literature?

I used EdgeR to get the list of DEGs (differentially expressed genes) from my liver RNA-seq experiment. I have done the gene overrepresentation and GSEA based pathway analysis. I am currently...

23 February 2021 8,969 1 View

Is it efficient to do liquid biopsy to investigate the gene expressions of the tumor?

To do liquid biopsy and look for specific gene expressions there are so many published papers. Thera are different protocols and procedures used to isolate circulating cell free RNA or mRNA from...

23 February 2021 7,326 5 View

How does the injection volume impact LLOQ and ULOQ in regression analysis (LC-MSMS)?

Hello, I have run into an issue that has sparkled a debate at work: is the LLOQ/ULOQ impacted by injection volume under the following circumstances: 1) Same dilution factor of all samples 2)...

23 February 2021 9,159 3 View

RNA degraded after good nanodrop readings?

23 February 2021 8,850 2 View