The Use of Steel-Tip Homogenizer Probe Results in Sample Carryover Contamination

The Use of Steel-Tip Homogenizer Probe
Results in Sample
Carryover Contamination

By Brian E. Mace and Patrick M. Sullivan,
Duke University Medical Center (Durham, North Carolina)
Introduction    Procedure     Results


In any experiment involving different treatment groups or varying starting material it is important to ensure that no crossover or contamination of sample material occurs. We routinely work with mouse models of varying genotype and then study the effects of different treatments in these mice. More specifically, we measure the levels of apolipoprotein E protein and mRNA in brain tissue using a very sensitive and quantitative assay system. Therefore, we tested two probes for potential contamination between tissue samples during the homogenization process. The probes we tested were the plastic-tip disposable generator probe 04727-50 and the steel-tip probe 04727-69.


All tissue was frozen in liquid nitrogen before homogenization. One half of the brain was homogenized with the plastic probe and the other half with the steel probe. The tissue was placed into 1 mL of guanidine buffer (5.0 M guanidine-HCL, 50 mM Tris pH 8.0) and allowed to thaw for 30 seconds before homogenizing on ice using PCR Homogenizer kit 04727-00. Each sample was homogenized for 45 seconds on low, medium, and high speeds in a 2-mL vial. The probes were then cleaned in water on high speed for two minutes, in homogenization buffer for 1 minute (twice), and wiped down with a Kimwipe (Kimberly-Clark). Homogenates were spun in an Eppendorf microcentrifuge at 14,000 rpm for 20 minutes at 4°C, and the supernatant assayed for protein content.

The micro BCA Protein Assay (Pierce) was used according to the manufacturer’s protocol. In brief, 150