mRNA down-regulated but protein in Western Blot still there. Which is the most plausible explanation?

How is it possible that RT-qPCR demonstrate a down-regulation of my interested gene (from 22 to 27 ct) while WB is showing the protein expressed with the same intensity as in cells where the gene...

26 September 2020 2,860 15 View

Mass Spec to identify proteins recruited in a functional regulatory region?

Hi everyone, I am trying to set up an experiment with the aim to identify putative transcription factors recruited in a functional regulatory region located in a non coding portion of human...

12 June 2019 7,499 2 View

Do you have any experience with HS578BST cell line?

Hi everyone, I am trying to do some experiments with HS578BST cell line (normal breast cell line) but I have big problems on growing them. I am using DMEM (with 10% FBS) and EGFR supplied every...

16 November 2018 9,218 3 View

Does T4 DNA ligase buffer interfire with Electroporation (nucleofector protocol) buffer?

Hi I am trying to electroporate a phosphorilate PCR product in my cell line. As the amount is very low, I was wondering if I can skip a purification step after phosphorilation. Does the T4 DNA...

17 October 2017 1,840 2 View

How can we measure teacher enthusiasm?

10 December 2013 5,853 9 View

Buffer to disrupt platelet alpha granules for PF4 quantification?

Hello! I want to quantify by ELISA the secreted (from platelets poor plasma) and the non-secreted (from platelet lysate) PF4 before and after TRAP stimulation. I will use the ELISA from R&D...

03 March 2021 1,499 2 View

Buffer Exchange using column packed with sephadex G-25 effect on protein concentration?

Hello, If i am doing a buffer exchange for an antibody of 1mg/mL, does the elution lose protein in the process of buffer exchange? For example, if i flow through 500 uL of 1mg/mL sample, and...

03 March 2021 6,299 3 View

How optimize the Size Exclusion Chromatography?

Hello! I'll do a size exclusion chromatography, but I only have an open column, and I'll perform the peptide extraction from yeast, using buffer lysis (sodium phosphate 50 mM/NaCl 30 mM/DNAse and...

01 March 2021 2,215 2 View

Why are my bands weird in this Agarose gel?

Can someone please give me some possible things that could go wrong? Here is my recipe: 0.5g Agarose 50 mL of TAE 1x 1 uL ethyl bromide. Gel was run at 100V for 1 hour. The buffer used is also TAE.

01 March 2021 9,952 3 View

What do I do with DLS peaks that are from the buffer?

I'm looking at the aggregation of my protein sample using DLS. Unfortunately, my buffer (20mM HEPES) also results in a set of peaks. These are at approximately 1 and 1000 d.nm. The lowest peak...

01 March 2021 9,015 2 View

What is the most suitable resuspension buffer for TCA/acetone precipitated protein?

I need to extract protein from fermented carbon sources for Bradford assay. Most researchers experiencing insolubility of pellet in resuspension buffer. Please assist me to select most suitable...

26 February 2021 7,129 3 View

What urea buffer can i use for biotagged protein extraction from streptavidin beads?

Hi all, I have been doing research on biotagged LDB1 proteins in Mel cells. I purify the proteins using M280 streptavidin beads. The yield is very low so I've been trying to optimize it by...

24 February 2021 5,749 3 View

Cloudy/high background western blot - can anyone help me?

Hi everyone! I try to detect VDR protein (~48 kDa) using Western Blot. After SDS-PAGE using 10% MP TGX Stain-Free Gel from BioRad I transferred the protein onto an metOH activated PVDF membrane...

23 February 2021 6,266 3 View

Have anyone checked before Cyclic voltammograms of intial ITO substrate and E-teck?

As a beginner in electrochemistry, I tried to obtain cyclic voltamograms of commercial anatase powder deposited on an ITO substrate or on Eteck. After several attempts, I noticed that my cyclic...

23 February 2021 3,175 3 View

Fictitious Boundary Meshing Method (FBM) in Commercial FEA software?

Is it possible to implement FBM in a commercial FEA software package, such as FEMLAB, COMSOL, or open source software like Moose?

22 February 2021 2,968 1 View