Finger-stick Blood Sampling Methodology for the Determination of Exercise-induced Lymphocyte Apoptosis

Summary

Exercise is capable of inducing apoptosis in immune cells. There are various measurement limitations, particularly relating to the amount of time required to isolate and treat a blood sample prior to the assessment. Demonstrated is a rapid and minimally invasive procedure for the analysis of exercise-induced lymphocyte apoptosis.

Abstract

Exercise is a physiological stimulus capable of inducing apoptosis in immune cells. To date, various limitations have been identified with the measurement of this phenomenon, particularly relating to the amount of time required to isolate and treat a blood sample prior to the assessment of cell death. Because of this, it is difficult to determine whether reported increases in immune cell apoptosis can be contributed to the actual effect of exercise on the system, or are a reflection of the time and processing necessary to eventually obtain this measurement. In this article we demonstrate a rapid and minimally invasive procedure for the analysis of exercise-induced lymphocyte apoptosis. Unlike other techniques, whole blood is added to an antibody panel immediately upon obtaining a sample. Following the incubation period, red blood cells are lysed and samples are ready to be analyzed. The use of a finger-stick sampling procedure reduces the volume of blood required, and minimizes the discomfort to subjects.

Protocol

1. Finger-Stick Blood Sampling

1. Collection of whole blood is typically taken at rest for a baseline measurement (usually following 10-15 min seated rest), during or immediately following an exercise bout, and throughout the post-exercise period (generally from 1-3 hours following cessation of the exercise).
2. Using Universal Precautions, first sterilize the fingertip with 70% isopropyl alcohol.
3. Next, puncture the site using an automated lancet or similar device. Although we use a 30-gauge lancet to provide for subject comfort, a larger gauge needle could be utilized.
4. Wipe away the first drop of blood, and then collect blood int....

Discussion

A minimally invasive sample collection procedure and subsequent analysis is demonstrated for the analysis of both early and late phases of exercise-induced lymphocyte apoptosis. It is also possible to clearly exclude those cells, which are necrotic, and not apoptotic. This procedure overcomes the drawbacks and discomfort associated with invasive venipuncture from the antecubital region at multiple time points prior to, during, and immediately after exercise, or securing a catheter for the duration of the exercise bout. I.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Flow Cytometry Binding Buffer		eBioscience	00-4222-26
RBC Lysis Buffer		eBioscience	00-4333-57
Lancet Device	Accu Check Soft Touch	Roche Group
Capillary tubes	Heparinized Capillary Tubes	Chase Scientific Glass, Inc.	501
Centrifuge	GmC Lab	Gilson, Inc.	PMS-880
Annexin V-FITC Conjugated		Biovision Inc.	K201-400-1
CD4-PE Conjugated	Anti-human, clone RPA-T4	eBioscience	12-0049-42
CD8-PE Conjugated	Anti-human, clone OKT8	eBioscience	12-0086-73
CD19-PE Conjugated	Anti-human, clone HIB19	eBioscience	12-0199-42
7-AAD Viability Staining Solution		eBioscience	00-6993-50
CD45RA-APC Conjugated	Anti-human, clone HI100	eBioscience	17-0458-42
Flow cytometer	C6	Accuri