Do you have any suggestions for neurosphere assay protocol, using adult mice?

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Does anyone have good protocol for dissociation of hippocampal neurons from adult mouse (flow cytometry/RNA seq)?

I am looking for any good protocol for dissociation of single hippocampal neurons from adult mice. We'll perform flow cytometry and eventually work on qPCR. As our lab is not specialized for...

06 July 2019 8,298 2 View

Are there any reliable antibodies for excitatory (glutamatergic) and inhibitory (GABAnergic) neurons in hippocampus?

I have Camk2a-Cre mice model, and would like to have antibodies for excitatory (glutamatergic) and inhibitory (GABAnergic) neurons in hippocampus CA1 regions. Do you know any good antibodies for...

10 November 2018 9,467 4 View

Any (hippocampal) culture and cell line useful to investigate epigenetic regulation (ie. high proliferation rate)?

I am currently looking for (preferably hippocampus) culture/neurosphere/cell line with high proliferation rate, which are available to purchase in Canada.

01 February 2018 4,687 2 View

Problems in using mouse host primary antibodies on mouse brain sample (IF/free floating staining).

I am doing free floating immunofluorescent staining on my mouse brain sample. I've been having difficulty using rodent host (ie. mouse, rat, guinea pig) primary antibodies, and I don't know why....

01 January 1970 6,925 13 View

Having difficulty IF staining on 5XFAD mouse brain sample (free floating).

I am not sure if we are supposed to change staining protocol for different mouse strains, but I am having difficulty staining (free floating immunofluorescent staining) on my 5XFAD mouse brain...

01 January 1970 3,227 5 View

Are there any rodent behavioural experiments that can substitute contextual fear conditioning test?

I am investigating the role of histone modification during hippocampal memory formation. It is the best if I could perform contextual fear conditioning on mice. However, we unfortunately do not...

01 January 1970 9,449 1 View

Reversal learning deficit, but normal memory extinction on Morris water maze test

I am wondering if anyone has seen the results of morris water maze like this; - normal acquisition (decrease in escape latency) BUT - significant deficit in reversal learning - normal memory...

01 January 1970 8,033 4 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...

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RIA. How to measure?

When you use RIA, with a control the unknown sample with antibodies, you bare in mind the binding sites of the antibodies. However you still need to measure labelled antigen (radioactive) and not...

28 February 2021 2,133 3 View

For gelatin coating for culturing HUVEC.Should the gelatin be made in sterile water or PBS? Do I need to autoclave the gelatin soln after making it ?

I want to use 0.1% gelatin. Should that be made in PBS or sterile water? Should I autoclave the gelatin soln after making it in water/PBS?

28 February 2021 274 2 View

The reason that the culture of white fungi Contamination with green mold(Trichoderma), the culture autoclave at121 for 15 min. What is it?

Is the period to autoclave not enough? The inoculation in a hood and flame and UV and alcohol.

27 February 2021 9,356 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

EMSA troubleshooting. Protein-DNA complex not migrating, cold probe lane with multiple bands. Can anyone help?

I'm running an EMSA to show the DNA binding activity of recombinant proteins versus the Wild type. Previous assays run by my research group using a different test system show the protein-DNA...

24 February 2021 8,325 1 View

David S. Thaler

PROTOCOL REVIEW

Eyes Wide Open: A Critical Review of Sphere-Formation as an Assay for Stem Cells

Erika Pastrana, Violeta Silva-Vargas, Fiona Doetsch📷

Nature Publishing Group

Cell Stem Cell Volume 8, Issue 5, p486–498, 6 May 2011

Open ArchivePlumX Metrics📷DOI: https://doi.org/10.1016/j.stem.2011.04.007

Cited ~500 times

Yuka Obayashi

Hello David, I am very sorry for delayed reply, but thank you very much for your suggestion! It was helpful!