Toward Zero Variance in Proteomics Sample Preparation: Positive-Pressure FASP in 96-Well Format (PF96) Enables Highly Reproducible, Time- and Cost-Efficient Analysis of Sample Cohorts
Stefan Loroch, Dominik Kopczynski, Adriana C. Schneider, Cornelia Schumbrutzki, Ingo Feldmann, Eleftherios Panagiotidis, Yvonne Reinders, Roman Sakson, Fiorella A. Solari, Alicia Vening, Frauke Swieringa, Johan W. M. Heemskerk, Maria Grandoch, Thomas Dandekar, Albert Sickmann
Abstract
= 0.9993, as demonstrated for mouse heart tissue lysate in 40 technical replicates. The targeted quantification of 16 peptides in the presence of stable-isotope-labeled reference peptides confirms that PF96 variance is barely assessable against technical variation from nanoLC-MS instrumentation. We further demonstrate that protein loads of 36-60 μg result in optimal peptide recovery, but lower amounts ≥3 μg can also be processed reproducibly. In summary, the reproducibility, simplicity, and economy of time provide PF96 a promising future in biomedical and clinical research.