Norbert Volkmar

11 Questions 10 Answers 0 Followers

Questions related from Norbert Volkmar

In vitro lipid / cholesterol delivery to membrane preparations?

Is there an easy way to deliver lipids in vitro to crude membrane preparations? The membranes have been prepared from cell lysates by ultracentrifugation in non-denaturing, detergent-free buffer....

16 November 2022 8,341 0 View

Sucrose gradients and mass spectrometry to analyse protein complexes?

I was wondering if anyone could relate their experience with isolating and analysing protein complexes in complex samples (whole-cell lysates) using mass spectrometry on sucrose gradient...

17 March 2022 10,105 5 View

Medium to grow HEK293 Freestyle in suspension as well as adherent?

Hi, can anyone recommend a cell culture medium that would allow me to grow HEK293 Freestyle cells in suspension as well as adherent cultures? Many thanks in advance!

07 March 2022 4,693 1 View

Why is sucrose added for the isolation of detergent-resistant proteins?

I noticed that in many protocols for the isolation of detergent-resistant proteins/aggregates sucrose is added either directly to the lysis buffer or the the lysate is centrifuged through a...

30 August 2021 7,767 1 View

How can I change the lipid composition of microsomes?

I am aiming to isolate microsomes from a cell line and then alter their lipid composition in vitro by artificially introducing lipids such as choesterol or phospholipids. The membrane proteins...

09 January 2021 5,952 1 View

Native protein extraction from cryopreserved, pulverised mouse tissue?

Can anyone recommend an efficient protocol for the extraction of native proteins from cryopreserved, pulverised mouse brain tissue? Ideally, this should also preserve membrane proteins. Thanks in...

31 August 2020 4,942 0 View

Has anyone done CRISPR/Cas9 in SGBS preadipocytes?

Has anyone done gene knockout using CRISPR/Cas9 in SGBS preadipocytes? Are these cells amenable to CRISPRX manipulation?

20 March 2019 7,418 1 View

Is there an easy way to identify lipid-binding domains in silico that are not necessarily trans-membrane helices?

I was wondering whether it is possible to extract some information concerning the propensity of hydrophobic stretches of sequences to interact with a particular type of lipids? The lipid binding...

17 September 2015 7,610 4 View

Mammalian expression vector for isolation of native protein complexes?

Hi, Can anyone recommend a specific mammalian expression vector that I could use to tag my protein of interest and that would allow me to purify any associated complexes for MS analysis? I am...

14 July 2014 5,559 10 View

Can the pcDNA5FRT/TO vector be used for transient overexpression of a gene of interest?

Hi, I have my gene of interest in pcDNA5/FRT/TO and would like to generate a stable cell line. Before generating the stable line, however, I would like to test expression by transient...

07 July 2014 270 3 View

Problem with Blue Native PAGE: Why is the transfer incomplete?

I am experiencing problems with one-dimensional Blue Native PAGE: It seems that proteins in the rage of 300 - 600 kDa are not properly transferred onto the PVDF membrane during Western Blotting of...

05 July 2014 9,551 5 View