Did anyone here ever did a successful Western blot for either Usp9x, USP7 or HUWE1 ?

02 June 2015 1 6K Report

I do semi-dry on PVDF, block with 4% milk and do the AB 1:1000 oN but all I get is background. Any help is appreciated!

Hi Inga,

I do semi dry transfer on PVDF but i use 5% milk for blocking.

To reduce the back ground I would suggest you to give more washes after primery incubation as well as after secondary antibody.

Try to change the primery Ab dilution.

Badges
Science method

More Inga Melzer's questions See All

What to do when you have reached the treshold of references in EndNote?

Dear fellows, yesterday, when trying to register another reference in EndNote, I got the message that the library was full. It has more than 5000 references. I wonder thus, what is the smartest...

25 June 2024 6,903 1 View

ANCOVA: how to control for change within control group?

In my study there are 2 intervention groups and 1 control group. In the ANCOVA I take trait scores as covariates and state scores as AV (so I compare the differences in the groups after the...

08 May 2023 2,967 1 View

When do people rather choose to travel by airplane than by car? How many hours of car driving at work ar professional employees willing to drive?

I am working on finding out the customer base who would be interested in taking the airplane between two Norwegian cities (Førde and Bergen). Driving by car takes at least 3 hours, and by bus...

12 March 2023 6,286 2 View

Why does my simulated flowout always look identical in QSWAT+ despite different simulation periods?

Dear all, I have come across a problem I have no explanation for. No matter which time period I simulate for my watershed in QSWAT+, the graph always looks identical. I have added a picture for...

07 March 2023 8,454 0 View

Experience with intestinal organoids in a organ-on-a-chip model?

I am looking for people who are also working with intestinal organoids (mouse or human) in an organ-on-a-chip system. I am working with the OrganoPlate from Mimetas and would be more than happy...

27 May 2021 3,108 0 View

From which day on does a chicken embryo feel pain?

Dear fellow colleagues :), I am looking for studies or sources on pain sensation in chicken embryos. What is known about the pain sensation of chicken embryos? From which day on can one assume...

16 June 2020 4,472 0 View

How do you teach your students clinical reasoning and how do you think it should be done in an ideal world?

We - an international consortium - are currently working on an EC-funded project to develop a longitudinal curriculum for clinical reasoning. Therefore, we are interested in how clinical reasoning...

11 March 2020 6,193 7 View

Could you suggest works dealing with the problems of contrastive theolinguistics?

I deal with the problems of contrastive theolinguistics, get acquainted with the works of colleagues available for reading. I would be grateful if you would point to scientific works in this area...

29 November 2019 3,282 3 View

How did the delta18O of the atmosphere look like in Earth's past?

Hi all, is there any information/paper if and possibly how the deltaO of the atmospheric oxygen has change in Earth's past? Thanks, Inga

29 August 2019 4,550 2 View

Statistical test for conditional regression?

Is there any statistical test or work-around to test whether the effect of variable a on c increases if b is high? All variables were asked for on a 7-point likert scale. I thought about...

17 July 2019 4,140 2 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 does my protein refolded to beta sheet during thermal denaturation analysis?

Hi! So i attempted to understand a novel protein behavior towards heat application by analyzing its secondary structure change. I subjected the protein to a thermal denaturation analysis using...

06 August 2024 1,989 3 View

Transfection in HEK293T cells?

Dear All, I am trying to transfect a pCDNA3.1 vector containing my gene of interest. The purpose is to figure out the localization of the protein of interest. I have fused the protein with GFP on...

31 July 2024 9,892 4 View

How to retain the a GFP tagged gene expression in stable cell line?

Hello , I established a stable cell line expressing GFP tagged to a centrosomal gene having G418 drug selection marker. I validated the stable line by IFA and Western blotting, results are fine....

29 July 2024 5,007 0 View

What is the relationship between protein structure and N or C terminal tagging choosing?

I want to do 2,3-butanediol dehydrogenase(BDH) enzyme purification to confirm its activity for 2,3-butanediol. Before that, I need to confirm which N or C terminal tagging is better for enzyme...

28 July 2024 366 3 View

Why cannot i find my protein on cell surface after antibiotic selection of expressing plasmid?

I cannot confirm cell surface protein expression by flow cytometry even after transfection and antibiotic selection of cells. Does it take long for proteins to express on the cell surface? the...

28 July 2024 3,178 3 View

EDC carbodiimide and NHS easter 's half life among different pH?

It has been long known that EDC carbodiimide and NHS easter's half life agains hydrolysis are highly related to pH. However, very surprisingly, not yet found a paper giving a table/chart of their...

25 July 2024 8,738 0 View

Does post-translational protein modification cause devisions on observed pI verses calculated pI?

In running two-dimensional gel electrophoresis on bacterial protein, some spots that appear to match a protein sequence have a significantly more acidic isoelectric point than the calculated pI....

24 July 2024 8,076 3 View

What is the function of fusion tag GAMGSGIQRPTST?

I found that several expression constructs of published protein structures containing an N-teiminal fusion tag GAMGSGIQRPTST. What's the founction of this fusion tag?

23 July 2024 1,353 4 View

The best source for amplification of ADAM17 prodomain?

hi every one I am making vector construction (for fusion proteins) and in this moment I wanna to amplification of ADAM17 prodomain with PCR. to yet, I couldn't amplified the ADAM17 prodomain with...

21 July 2024 8,660 1 View