The HeLa cells were fixed with 4% paraformaldehyde, and permeabilized with 0.1% Triton X-100. Then, the specimen was blocked in BSA/PBS blocking buffer. Subsequently, the specimen was incubated with Mouse Anti β-Tubulin IgG, and, after washing, was further incubated with biotin-conjugated Goat Anti-Mouse IgG Biotin Conjugate. Finally, it was rinsed with PBS and stained with Streptavidin FITC Conjugate at 5 μg/mL (shown in green) and DAPI-2HCl at 1 μg/mL (shown in blue)
Usually ships within 24 hours.
The fluorescein labeling kits are high-performance derivatives of fluorescein dye, activated for easy and reliable labeling of antibodies, proteins and other molecules for use as fluorescent probes.
Fluorescein dyes are used in wide-ranging applications including fluorescence microscopy, flow cytometry and immunofluorescence-based assays such as Western Blotting and ELISA. Fluorescein NHS are reactive toward primary amine groups on proteins, peptides and other biomolecules, while fluorescein-maleimide reacts with free sulfhydryls on cysteine residues.
Most activated fluorescein derivatives are mixtures of isomers with reactive groups attached at the 5- and 6-positions of the bottom ring. There is a big difference in flow cytometry although it is a small difference in the properties of these isomers in terms of excitation and emission spectra. The small difference will make your assay complicated because the ratio of two isomers varies lot by lot.
BroadPharm fluorescein labeling kits use single isomer- 6-isomer NHS and Maleimide to solve the issues.
* single isomer, no lot-to-lot variability.
Fluorescein is mainly used to label primary antibody and secondary antibody for the applications below:
CWR22R cells were fixed with 4% paraformaldehyde, blocked by 3% BSA. Subsequently the specimen was cultured at 4°C overnight with Anti PSMA monoclonal antibodies (1 : 100), and incubated with FITC-conjugated anti mouse IgG antibody at room temperature for 2h. DAPI nucleic stain was performed in blue.