Does anyone know how to use image segmentation in image J?

Hi, Im trying to use the image segmentation tool in image J for my image and Iam not getting it correctly to find out the co-localization of two proteins. If anyone has used it, could you please...

09 October 2019 6,877 4 View

Anone facing phototoxicity in IF samples?

Hi, Iam doing IF in HeLa cells and facing a weird problem. I use the standard protocol and when I expose the cell to laser light in the 488 channel, all the fluorescence fades the instant its...

08 September 2018 6,532 4 View

How to design a blocking peptide?

Hi, I want to design a blocking peptide for the C terminus of my protein. If anyone knows how to do it, please help me out. Thanks jaya

04 May 2018 3,735 3 View

Anyone working with connexins?

Hi, Iam encountering a problem in the cell culture. I have hela cells stably expressing connexin 36 attached to GFP on the C terminus. My problem is there is connexin 36 in the internal vesicles,...

11 December 2017 3,842 3 View

Help with cleaving GFP from the protein of interest ?

Hi, Iam working with a protein which is attached to GFP and expressed in Hela cells. I want to do single particle imaging using cryoEM. Im trying to do an IP with GFP antibody and trying to use a...

09 October 2017 2,326 1 View

How to represent western blot for paper ?

Hi, I am trying to represent my western blots for paper and have a clarification how to represent the Y axis in the graph. I normalized the test protein with actin and then divided my test protein...

08 September 2017 6,061 5 View

Is anyone working with halotag proteins?

hi, Iam working with a halotag attached to my protein expressed in Hela cells. I was trying to pull down the halotag with magne beads and got nothing in the western. I tried increasing the...

07 August 2017 8,554 3 View

Help with western blot. hello any one there?

hi, I am working with GFP tagged connexin 36 transfected in hela cells. I do a crude membrane preparation and then used chaps to solubliize the membrane for two hrs and run it on a 10% gel. When I...

02 March 2017 9,048 10 View

Should I give the SD in the fold difference and the P value for western blot for publicaiton?

Hi, Im confused on how to present my result for western blot for the paper. I did the experiments thrice by licor and quantified and for each time the machine gave me different values for the same...

09 October 2016 3,329 8 View

How much of cDNA to use for qPCR?

02 March 2016 375 5 View

Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence?

I am on the lookout for the Enhanced Yellow Fluorescent Protein (Aequorea victoria) DNA sequence. Does anyone know where I can find it? Thank you in advance

03 March 2021 3,568 1 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

POlymer+DMSO solution and moisture??

What could be the reason of moisture at the sides of Vial when I am preparing my polmyer+DMSO solution for membrane fabrication? What etiquettes I am possibly missing here?

28 February 2021 6,516 5 View

How can I represent SDS-PAGE results for machine learning analysis?

I am trying to classify and analyze the results of an SDS-PAGE based array for bacterial detection using machine learning, but I have trouble finding the best way to represent the results with...

27 February 2021 9,176 3 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

Why my agarose gel staining sometimes looks not homogenic but like gradient (see the figure)?

I am worried about this overexposure of the upper part of the gel in the picture. Is it possible that this is the result of too much concentration of ethidium bromide? Why is there such a big...

25 February 2021 8,140 3 View