Protective Immunity Project

Chris Larsen, MD, DPhil, PI

Christine Martens, Ph.D. Project Manager

Vicki Hertzberg, Ph.D. Director of Data Management and Analysis Core

3 studies

• 2 human– New renal transplants (n=60) and 20

controls, followed for 2 years– Existing renal transplants (n=60) and 30

controls, receiving flu vaccine, followed for short time

• 1 macaque study, 30 macaques randomized to 6 groups

Longitudinal studies

ID Time 1 Time 2 … Time t

1

…

n

Measurements

At each time point for each subject obtain measurements from:– Flow cytometry– ELISPOT/ELISA– M-array

Flow cytometry

For each subject at each time, take several blood samples.

For each blood sample, label all of the PBMCs with 8-10 markers (a panel). Cells will fluoresce according to the amount of each marker present.

Apply 6-8 panels for each subject at each time.

Each panel usually has 2 markers that are common to all other panels

What to the data look like?

• X-Y graph for fluoresence intensity for marker X vs marker Y.

• Gates are drawn that – Separate present from absent– Isolate cells of different types

Issues?

• How to analyze?

• Can we deal with fluorescence intensity rather than presence or absence?

• How do we institute QC for the gating?