DNA fluorescent staining · Principle: Detection of mycoplasma contamination using a fluorescent dye (bisbenzimide, Hoechst 33258). This dye binds to the Adenosine-Thymidine (AT) rich region of DNA. Because the DNA content of mycoplasma is predominant (55-80%), it can be stained and detected. After the cells contaminated with mycoplasma are stained, many small and uniform fluorescent spots can be seen outside the nucleus and around the cells, which is the DNA of mycoplasma, which proves the contamination of mycoplasma. (A) Negative result (Positive Result Blue fluorescent dots and filiform dots appear on the surface of the Vero cell and on the cell surface, indicating that the test sample is contaminated with mycoplasma. Onlyrelx Energy5000 disposable vape pen is portable and fashion disposable electronic cigarette, disposable ecigs pen are trending featured vape pen for vapors as it's safety and easy to use. Disposable vape pod,disposable vape, wholesale vape,vape wholesale,vape pen manufacturer and supplier.disposable vape pen,disposable electronic cigarette,disposable ecigs pen,disposable ecigs stick,disposable e-cigs pen,disposable vape factory,disposable vape pod,disposable vape device,vape pen,vape stick, vape wholesale,wholesale vape,customized dispsoable vape pen,customized vape pen,OEM&ODM disposable ecigs pen,disposable electronic cigarette wholesale, wholesale disposable electronic cigarette,distribute vape pen,vape pen distribute,high quality vape pen,high quality vape pod. Onlyrelx Energy5000,Customized Vape Brand,Customized Vape Pen,Custom Made Vape Pen Shenzhen Onlyrelx Technology Co.,Ltd , https://www.onlyrelxtech.com
· Features: Simple, economical and sensitive, widely used as a routine detection step. It is possible to detect mycoplasma that is difficult to culture, such as M. hyorhinis, which is faster than direct culture and can be known in about a week.
· Disadvantages: Sometimes there will still be a fluorescent background that will affect the interpretation.
step:
• Remove the cultured Vero cells and aspirate the culture. 2 ml of 1:3 mixed glacial acetic acid/methanol fixing solution was added to each well, and after standing for 10 min, the fixing solution was aspirated and washed three times with PBS.
· Add 1 ml of Hoechst 33258 working solution to each well and let stand for 5-10 min at room temperature.
· After aspirating Hoechst 33258 stain, wash it 3 times with sterile water, air dry, add 1 drop of sealer, and cover with a cover slip.
· Observed with a 100x-400x fluorescent microscope. Observe whether there are blue fluorescent dots or fluorescent spots outside the nucleus.
Result interpretation:
If there is mycoplasma contamination, blue fluorescent dots or filiform dots appear on the surface of the Vero cell and on the cell surface. It has the same shape and is different from the irregular spots in which the cell debris is dyed. Its fluorescence can last for weeks.
Legend (A) is negative result: Under the fluorescence microscope, only the nuclei of Vero cells were observed, and the test samples were free of mycoplasma contamination. (For positive result: Blue fluorescent dots and filiform dots appear on the surface of the Vero cell and on the cell surface, indicating that the test sample is contaminated with mycoplasma.
Under the fluorescence microscope, only the nuclei of Vero cells were observed, and the test samples were free of mycoplasma contamination.