high fluorescence values

Dear all,

 

I am having a problem with high fluorescence values in some of my
experiments. I basically stain human PBMC for different markers, looking at
monocytes, pDCs and mDCs. I acquire in a FacsAria doing automatic
compensation using BD Compbeads. My unstained beads are correctly placed
under 10E1 for each channel. However, on the experiments, it is quite usual
to have values for the negative cells that are around 10E3. (Could this be
due to autofluorescence?, it does seem to be higher in APCs than in T
cells). This phenomenon is independent on the fluorochrome being used. I’m
still able to clearly see my positive population which is even more
fluorescent, so it doesn’t worry me that much… (should I start worrying????)

Any insight into this will be very much appreciated.

Thanks! 

Candela

 

María Candela Iglesias, PhD

Centro de Investigacion en Enfermedades Infecciosas (CIENI)

Instituto Nacional de Enfermedades Respiratorias

Calzada de Tlalpan 4502. Col. Seccion XVI

CP 14080. Deleg. Tlalpan

México DF.

tel y fax +52 (55) 56 66 79 85

candela@iner.gob.mx

candela.iglesias@cieni.org.mx