How to prepare FITC solution?
Dissolve the FITC in anhydrous DMSO at 1 mg/ml. Note: This should be prepared fresh for each labeling reaction. 3. For each 1 ml of protein solution, add 50 µl of FITC solution, very slowly in 5 µl aliquots while gently and continuously stirring the protein solution.
What color is FITC?
FITC exhibits an excitation maximum at λ = 495 nm and emission maximum at approximately λ = 519 nm. The color of the compound is yellow while the emitted light is green.
What is FITC stain?
Fluorescein-5-isothiocyanate (FITC) is a fluorescence dye and belongs to the xanthene dyes. FITC is used for labeling of different biomolecules, e.g. immunoglobulins, lectins and other proteins, peptides, nucleic acids, nucleotides; oligo-and polysaccha- rides.
What is degree of labeling?
The Degree of Labeling (DOL), sometimes called Degree of Substitution (DOS), is a particularly useful parameter for characterizing and optimizing bioconjugates, such as fluorophore-labeled proteins. It is expressed as a molar ratio in the form of label/protein.
What is f p ratio?
In flow cytometry, the effective F/P ratio is the average apparent number of fluorochrome molecules conjugated per primary antibody, as determined through fluorescence measurements taken on the flow cytometer.
What is the molar ratio between oxygen and water?
The mole ratio between oxygen and water is 1:2.
How to stain phalloidin FITC methanolic stock solution?
When staining, dilute 5 μL Phalloidin-FITC methanolic stock solution into 200 μL PBS for each coverslip. To reduce nonspecific background staining add 1% bovine serum albumin (BSA).
How do you stain in vitro stimulated cells?
If using in vitro stimulated cells, simply resuspend previously activated cultures in Cell Staining Buffer and proceed to Step 2. Add Cell Staining Buffer up to ~15ml and centrifuge at 350xg for 5 minutes, discard supernatant.
What is the protocol for immunofluorescence staining?
Immunofluorescence Staining Protocol (from IHC world) 1. Preparation of Slides A. Cell Lines Grow cultured cells on sterile glass cover slips or slides overnight at 37 º C Wash briefly with PBS Fix as desired. Possible procedures include:
What kind of staining is used for apoptosis?
Cells that have lost membrane integrity will show red staining (PI) throughout the nucleus and a halo of green staining (FITC) on the cell surface (plasma membrane). The above is an example protocol for PS exposure detection using annexin V, based on the protocol provided in Annexin V-FITC Apoptosis Detection Kit (ab14085).