06 June 2019 2 4K Report

I routinely functionally assess the iPSC derived neurons using calcium imaging with Fluo-4/AM (490nm/510nm). However, have not been successful at using those same neuronal dishes for carrying out ICC. The reason that I think could be are as follows:

  • The calcium-sensitive dye fluo-4/AM has an emission at 510nm. As a reason why, there is a spectral overlap with my anti-bodies of interest.
  • As I try and wash out the dye from the dishes, the neurons detach as a matter of fact. These neurons have been subjected to constant washing before and after the calcium dye-loading process. Moreover, these neurons are first stimulated with TTX following ionomycin and EGTA+TX are added to the dishes as internal controls.

    • Any suggestions or references or direction to a protocol would be of a great help!

    Thanks!

    More Yojet Sharma's questions See All
    Can addition of GABA agonist block transients in neurons(See description for details)?

    I am trying to characterise GABA developmental switch in iPSC derived neurons using Ca2+ imaging. However, as I add GABA agonist, I observe the spontaneous activity of several neurons being...

    08 September 2019 694 2 View

    What markers can be used to distinguish neural progenitor cell from a neural stem cell?

    How can we asses if the population of cells in culture are neural stem cells or neural progenitor cells? Will progenitor cells also undergo neurosphere formation assay? Is there a way to identify...

    03 April 2018 5,588 1 View

    How do I confirm whether Tetrodotoxin citrate is working or not?

    I have TTX citrate ordered from Hello Bio. On using it at 1uM as my working concentration, it didn't show any effect. How do I test whether the toxin is actually working or not?

    01 February 2018 8,541 6 View

    What is the chemistry of calcium dyes?

    01 February 2017 2,147 8 View

    What imaging setup can be useful for calcium imaging?

    01 February 2017 3,517 1 View

    Similar questions and discussions
    Why does blocking with my immunogenic peptide cause nuclear fluorescence in ICC?

    Hi all, I am validating a polyclonal antibody raised against human ANT4, through WB and ICC. In most of my controls, the antibody shows very little off-target binding, but when I pre-incubate the...

    03 March 2021 5,999 3 View

    What is the production process of CaFe ferroalloys?

    I looking for ferro calcium production process

    03 March 2021 1,755 4 View

    No title

    02 March 2021 3,060 3 View

    12% ledeburitic cold work tool steel, etched for revealing retained austenite, any help for clarifying?

    Hello everyone. Given microstructure image belongs to a 12% ledeburitic cold work tool steel (annealed and tempered). The sample was etched with picral (100 mL ethanol and 4 g picric acid). I...

    02 March 2021 4,119 3 View

    Does Napabucasin soluble in dichloromethane and in ethanol?

    I need to prepare a solution of Napabucasin (inhibitor of cancer cell stemness) in dichloromethane. The drug is expensive, so maybe somebody already tried to do it. If it is soluble, what is the...

    02 March 2021 9,945 3 View

    New Journal, "Integrative Psychological and Behavioral Science" -- do you not know, and have you not seen, this done before?

    Question to you and THEM, the New Journal, "Integrative Psychological and Behavioral Science" -- do you not know, and have you not seen, this done before? There appears to be a core problem for...

    02 March 2021 3,024 2 View

    Help request in CST microwave?

    im working now on TPV solar cells and metamaterials and i realy need our help in CST microwave i have a tpv cell simulation and i want to extract particullarly the absorbtion of only the active...

    02 March 2021 2,246 2 View

    Do you know any MR image dataset?

    I would like to research on MR images (0.5T and 3T). Can you please suggest some websites that I can download dataset including both 0.5T and 3T MR images? Thank you.

    02 March 2021 7,735 3 View

    Does anyone know HRM channel in a 2 plex QIAGEN Rotor- Gene Q qPCR can be used as a separate channel?

    I am going to have 3 different probes in my qPCR work that I am going to do. But I realized that the machine we have in the lab is a Rotor-Gene Q 2plex HRM Platform, saying it has green, yellow,...

    01 March 2021 8,544 1 View

    Issues with Permeabilization during Flow Cytometry?

    Hello Everyone. Currently I am working to characterize macrophages in the myocardium after ischemia-reperfusion injury in rats. Due to the low total cell number isolated from rat hearts I can...

    01 March 2021 3,867 3 View

    Our partners will collect data and use cookies for ad personalization and measurement. Learn how we and our ad partner Google, collect and use data. Agree & close