14 Questions 12 Answers 0 Followers
Questions related from Martina Boncaldo
I have to perform WB on FFPE tissue. I've already done several attempts and the result is always the same. Can you help me understand why the bands are like this and not neat? Standard is fine (I...
23 June 2021 4,340 4 View
I have to perform WB on FFPE tissue. Do you know if there is a way to understand exactly how much buffer I should put in the sample after deparaffination (for instance, depending on the...
12 June 2021 5,668 2 View
I have a protocol that uses SDS (2%), tris-HCl (200 mM, pH 8), DTT (200 mM) and glycerol (20%) for the extraction of protein from FFPE tissue. Is there a way to understand how much...
12 June 2021 1,162 1 View
It's just a curiosity that could also help me in my troubleshooting: does Laemmli buffer behave "normally" without the lysate? Like, what happens if I load in the well 20 μL of Laemmli instead of...
27 May 2021 1,166 2 View
I have to perform Western Blot on FFPE tissuse First I did deparaffination, then I added the lysis buffer (20 mM Tris-HCl pH 8, 2% SDS) and I incubated for 20 minutes at 99°C and for 2h at...
27 May 2021 4,485 6 View
I have to perform Western Blot on FFPE tissue and the first part is the extraction. First I did deparaffination, then I added the extraction buffer (20 mM Tris-HCl pH 8, 2% SDS) and I incubated...
11 May 2021 8,821 4 View
It's a buffer for protein extraction from FFPE tissue, but it's not actually important for the answer. I just want to understand what to do exactly I want to prepare 100 mL of this buffer: 2%...
07 May 2021 442 3 View
I need to know different ways to extract protein from FFPE tissue without a kit. I already know how to do deparaffination, but I just know one buffer (Tris-HCl, DTT and SDS). Do you know other...
05 May 2021 3,690 1 View
I have to perform WB on FFPE tissue, but the protocol of the extraction from Bio-Rad is for cell culture. Do you know a protocol that involves RIPA buffer that I could use on FFPE tissue? Thanks...
05 May 2021 4,580 1 View
I used DTT in protein extraction for WB, but unfortunately it was stored at RT when in the label the storage temperature was 2-6°C. What are the consequences? What is DTT's role in protein...
04 May 2021 5,939 3 View
I have to perform WB on FFPE tissue, so I used a protocol to extract the proteins but after the electrophoresis I got only one band. This is the protocol I used Extraction buffer (1 mL): DTT...
04 May 2021 1,898 7 View
I tried to use an expired gel from Bio-Rad. It was stored in the fridge at about 3°C for several years. So what could change in the gel? Is it possibile that the pores of the gel got larger and so...
04 May 2021 1,283 1 View
I already have a protocol for deparaffinization, but I have doubts about the extraction buffer. I found a protocol which uses just three reagents (SDS, Tris-HCl and DTT), but I have to use a...
26 April 2021 3,189 2 View
I have to perform western blot on ffpe tissue, but in the official protocol of the material that I use in lab there's just a method for fresh/frozen tissue. Moreover in majority of web reviews the...
26 April 2021 3,609 0 View
