Weird Black Line on Western blot ?

More Saverio Charalambous's questions See All

Why does my PCR amplification produces non-specific bands in a consistent manner?

Hello, I am running a PCR amplification using NEB Q5 high fidelity polymerase kit on 1.4-1.6 kb gBlock fragments with the following reaction conditions: > 30s at 98°C > 10s at 98°C -...

28 July 2019 1,990 4 View

Weird Black Line on Western blot ?

I have some gels that have a consistent black line at unexpected protein size consistent throughout gel? What could be the reason? Thank you

06 July 2019 6,532 3 View

Why does my PCR amplification produces non-specific bands in a consistent manner?

Hello, I am running a PCR amplification using NEB Q5 high fidelity polymerase kit on 1.4-1.6 kb gBlock fragments with the following reaction conditions: > 30s at 98°C > 10s at 98°C -...

06 July 2019 9,650 4 View

Does anyone know what might be causing the smeared bands on my western blot?

I got these smeared bands quite often lately. We typically run the gel at 140V with a 10-12% gel and do a wet transfer at 220 mA for 1.5 hr in cold room. We also noticed some dirty spots/dots (see...

10 August 2024 7,480 3 View

Why is my ASO degraded quickly on agarose gel?

I have been working on Red blood cell-derived extracellular vesicles as Antisense Oligonucleotide (ASO) carriers. We normally run agarose gel to quantify the loading efficiency. I used naked ASO...

06 August 2024 3,130 2 View

Low-yield gel extraction problem?

I am having an issue with my gel image where my PCR product is not appearing very bright on the gel. When I perform gel extraction, the A260/280 purity value is very low. I used the Qiagen gel...

05 August 2024 9,798 3 View

PCR showing no bands - Master mix and primers didn't mix?

Hello everyone, I performed a PCR yesterday, and the results showed no bands on the gel. Of course, I probably missed some crucial steps, like adding my samples to the PCR strips themselves, for...

31 July 2024 2,406 6 View

How to check pH of a high strength hydrogel?

We are working on biopolymeric hydrogels. Our system is highly viscous and sticky, and the gel formed are high in strength. We are unable to use pH electrode and pH strip. Please suggest an easy...

30 July 2024 942 2 View

Following click reaction in cell lysates, protein is immobile and remains at the top of the gel in SDS-PAGE?

I am using CuBr/THPTA for a click reaction in total cell lysates. I am facing issues with my protein sample in non-reducing SDS-PAGE where it's not migrating properly and most of it remains at the...

29 July 2024 950 4 View

Why my IgG antibody looks fragmented on non-reducing SDS-PAGE?

I am performing IgG purification and I have to show my results on SDS-PAGE. I use 10% tris glycine gel and prepare the samples under non reducing conditions. I am new to antibodies and therefore...

23 July 2024 6,664 6 View

What is the cause of this aggregation just bellow 70 kDa mark?

Hello, I was running a 12% SDS Page electrophoresis on few granulosa cell samples and got this result after the ponceau staining. The total protein lysate seem to aggregate at 70 kDa ladder mark...

21 July 2024 5,128 4 View

Why my gel electrophoresis have shadow bands? Please see the attached picture for the gel electrophoresis ?

Sometimes I see the shadow like bands and its not true band. I want to know that what's the reason for it. I am using 2% gel for running genotyping samples I have uploaded the gel picture in both...

19 July 2024 148 6 View

What will be the best way to test NFkb activation via western blot?

Hi, I have lysate from human macrophages, and I wonder what will be the best way to check the NFKB activation in those cells. I was thinking p-p65 or p-ikkb./ Is that make sense?

18 July 2024 5,214 4 View

Pierre Béguin

It looks like there is some contaminant in the electrophoresis buffer, hence its appearance across the whole width of the gel

Saverio Charalambous

Thank you Pierre Béguin ! Would you recommend changing or using newly made running buffer?

I would definitely recommend to make fresh running buffer