Currently I am working in identifying calcium profiles from two mice lines a WT and a Mutant, and I would like to use this specific assay to observe any differences between them.
As I was reading some published research references, they'd used the e. coli supernatant after stimulating the macrophages for 24hrs with LPS. However, I would be interested in an assay in which it is possible to add the bacteria directly to the cells after the LPS stimulation while running the calcium assay. I was wondering how many CFU/mL of bacteria would be ideal to use to get a nice stimulation in macrophages.
I using Fura2-AM and Flou4 in my assays.
Hi. first of all, forgive me if I'm not answering your question right.
previously, I want to determined the production of NO in the macrophages in two different group (infected and non infected), after treatment with certain active compounds. For the infected group, the MOI (multiplicity of infection) relies on the number of cells that you want to seed in each flask/ well. After calculation, the bacteria inoculum was introduced into the cells directly, incubated for 2 hours and followed by treatment with compound for overnight. Next, the cells were harvested and I used the supernatant for determining the NO. I did use LPS +bacteria+ compound + cells as a control.
Sorry if I'm not answering your question.
- Badges
- Science topic
- Similar topics
- Biological Science
- Microbiology
- Microorganisms
- Bacteria