38 Questions 12 Answers 0 Followers
Questions related from Amanda Rocha
I need quantify the protein amout from human DCs lysed by RIPA buffer ( 150 mM NaCl, 1% NP-40 or Triton X-100, 0.5% sodium deoxycholate, 0.1% SDS, 50 mM Tris, pH 8.0 ) to use in a immunoblot....
12 December 2019 3,868 3 View
I need to detect and measure the abundance of cytoplasmatic proteins (CASP1 an GSDMD) by western blot in human mo-DCs, and some companies where I purchased the antibodies do not specify a...
05 May 2019 1,505 5 View
I have purchased THP-1 cell line from ATCC and they recommed mercaptoethanol. It is a reducing agent, what is the function of this kind of agent in culture medium of THP-1 cells? How...
05 May 2019 10,247 3 View
I work with human mo-DCs and I need to detect cytoplasmatic and nuclear proteins by flow cytometry using flurochrome-conjugated antibodies (direct detection), that is, I need to permeabilize both...
05 May 2019 5,085 5 View
Is it best perform fixation step with formaldeihyde or paraformaldeihyde at 4ºC or room temperature or there is no great difference? I work with human mo-DCs in culture and I need to detect...
05 May 2019 3,527 4 View
I need to detect and measure the abundance of cytoplasmatic proteins (AIM2, IFI16, GBP1, GBP4) using direct detection by flow cytometry with fluorochrome-conjugated antibodies in human mo-DCs and...
05 May 2019 6,702 4 View
I need to detect and measure the abundance of cytoplasmatic proteins (AIM2, GBP1, GBP4) and nucleic proteins (IFI16), all apparently soluble, using direct detection by flow cytometry with...
05 May 2019 5,731 3 View
I need to detect and measure the abundance of cytoplasmatic proteins (AIM2, GBP1, GBP4) and some nucleic proteins (IFI16)using direct detection by flow cytometry with fluorochrome-conjugated...
05 May 2019 7,690 2 View
Someone that work with human primary cells and western blot: how many cells I need to identify protein abundance in western blot analysis? I want to quantify Caspase-1 and gasdermin D
01 January 2019 5,241 5 View
I have pX330 available in my lab, but I haven't found many recent works using this plasmid.
11 November 2018 1,317 3 View
Someone have ever tried to differentiate human DCs from cell line that resemble mo-DCs?
11 November 2018 6,340 5 View
IL-1B/IL-18 secretion, westen blot analysis of 2 subunits of caspase-1, FAM-FLICA detection or another method?
11 November 2018 2,548 2 View
Or it depend on the kind of plasmid used?
11 November 2018 1,007 2 View
I need a positive control to my caspase-1 activation assays in human monocytes-derived dendritic cells in culture.
11 November 2018 7,571 3 View
What is the best brand of antibodies to detect human proteins through WB and/or flow cytometry?
11 November 2018 463 4 View
I need to analyze a specific intracellular protein level in control and infected human monocyte-derived dendritic cells
11 November 2018 6,757 5 View
I want to detect caspase 1 activation in human monocytes-derived dendritic cells after interaction with my pathogen.
11 November 2018 2,203 4 View
How can I detect pyroptosis in human mo-DCs induced by a pathogen? Are there some comercial kit?
11 November 2018 5,229 0 View
I have analyzed mRNA human samples in bioanalyzer and the RNA concentration provided did not match with the RNA concentration of the same sample in the same dilution provided by Qubit. What RNA...
05 May 2018 7,674 3 View
Someone already tried to perform qPCR with a multichannel pipette? Or is it not recommended?
05 May 2018 7,248 6 View
I need to perform qPCR of 40 human genes and I would like to know if I can purchase primers that already have tested and have confirmed efficiency of >90-95%
03 March 2018 4,470 4 View
I’ve been performing, so far, my qPCRs reactions in 50 uL of final volume; however, at doing in this way, I have spent a lot of material. In this sense, I would like to know if I could perform my...
03 March 2018 9,180 4 View
If I switched the final volume of my qPCR reactions from 50uL to 25uL, will it interfere in my analyzes of my biological replicates, once I already started the analyzes of some replicates with 50...
03 March 2018 712 4 View
I am trying to define a standard curve for the primers that I have designed for qPCR and I ordered to sythetize the correpondent oligo for test the primers in different concentrations, but I don't...
11 November 2017 7,966 2 View
How much I can rise or decrease temperature without compromise RNA integrity?
08 August 2017 2,356 3 View
DNAse protocol that I am using in my RNA samples says to inactivate DNAse using phenol-cloroform and shows a complex and prolonged protocol to inactivate. There is an alternative method that I can...
08 August 2017 9,184 3 View
What's the method used to measure the RNA integrity?
05 May 2017 8,234 2 View
Does anyone can explain why I need to use A 260/280 to measure RNA amount? What this means?
05 May 2017 6,807 4 View
How bioanalyzer determine the RIN of RNA sample? Why in some cases the RIN appears as not applicable? What could happened?
05 May 2017 3,385 4 View
How method I can use to obtain dendritic cells from monocytes? Or there is a better way to obtain high amount of these cells?
01 January 2017 523 2 View
What's the major difference about using these two kits for quantify cytokines in supernatant of culture cells? Both have the same aims?
01 January 2017 5,960 2 View
I have co-cultured dendritic cells with T. cruzi and I need to evaluate the cytokines realized from this interaction. How can I harvest and storage the supernatant after the infection to perform...
01 January 2017 3,428 12 View
I would like to know what cytokines are produced by dendritic cells. I need to know ,specifically: These cells can produce IL-1beta, IL-6, IL-10, IL-12p40, TGF-beta, and TNF-alfa?
01 January 2017 2,357 4 View
I need do quantify the cytokines that are produced from dendritic cells descendant from monocytes when I infect these DCs with Trypanosoma cruzi in a 24-well-plate, but I have never worked with...
12 December 2016 8,050 11 View
Could someone that work with human mo-DCs transfection with siRNAs recommend a best method to do this? I generally have seen works using lipofectamine 2000 (Invitrogen), eletroporation (Amaxa/bio...
01 January 1970 3,326 3 View
I have performed a antibody titration to detect a specific cytoplasmatic protein in human monocytes starting with the double amount of general antibody recommendation (2 ug) and I have made 9...
01 January 1970 1,640 0 View
I have observed that both index are used in order to evaluate the best concentration of antibody in FC, but which one is most recommended to discover the best antibody concentration that provide...
01 January 1970 3,969 4 View
Someone that work with human dendritic cells and western blot: how many cells I need to identify protein abundance in western blot analysis? I want to quantify Caspase-1 and gasdermin D
01 January 1970 2,526 4 View
