BD fastimmune protocol

Calman Prussin CPRUSSIN at
Wed Feb 26 19:41:13 EST 1997

I am concerned that by only collecting 3000 CD3+ cells you will be 
generating less than the most robust numbers for IL-4 frequencies. The 
significance of a frequency is proportional to the number of events 
you acquire. David Parks has a table in the FACS chapter he wrote for 
Paul's Fundamental Immunology, 2nd edition. By only collecting 3000 
events, a value of 5% could be anything from 4 to 6%. By increasing 
that to 50,000 events, you narrow the range to 4.7% to 5.3%.

Calman Prussin
Laboratory of Allergic Diseases
NIAID/ National Institutes of Health

From: 	Michelle N Fiordalisi
Sent: 	Wednesday, February 26, 1997 9:39 AM
To: cyto-inbox
Subject: 	Re: BD fastimmune protocol

Since I've had several requests for a workable protocol for 
cytoking staining, I'd lot I'd post it for every one.

The procedure I've been using is based on the BD protocol published in 
their Application Note 1.

[text deleted]

I then acquire 3000-5000 events in a CD3+ gate and analyze by
quadrant analysis (CellQuest) of dot plots.  I calculate % positivity 
using BD's formula:  (Activ. sample - activ. isotype control) - ( 
sample - Unstim. isotype control).

-- Michelle

Michelle N. Fiordalisi, Ph.D.
Clinical Immunology Fellow
University of North Carolina Hospitals
email: mnfiord at
Phone: 919-966-4058  FAX: 919-966-0486

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