Microfluidic Leukocyte Isolation for Gene Expression Analysis in Critically Ill Hospitalized Patients

• Published in: Clinical chemistry (Baltimore, Md.) Vol. 54; no. 5; pp. 891 - 900
• Main Authors: Russom, Aman, Sethu, Palaniappan, Irimia, Daniel, Mindrinos, Michael N, Calvano, Steve E, Garcia, Iris, Finnerty, Celeste, Tannahill, Cynthia, Abouhamze, Amer, Wilhelmy, Julie, Lopez, M. Cecilia, Baker, Henry V, Herndon, David N, Lowry, Stephen F, Maier, Ronald V, Davis, Ronald W, Moldawer, Lyle L, Tompkins, Ronald G, Toner, Mehmet, Inflammation and Host Response to Injury Large Scale Collaborative Research Program
• Format: Journal Article
• Language: English
• Published: Washington, DC Am Assoc Clin Chem 01.05.2008; American Association for Clinical Chemistry; Oxford University Press
• Subjects: Analytical, structural and metabolic biochemistry; Arrays; Biological and medical sciences; Blood; Burns - pathology; Cell Separation; Critical care; Critical Illness; E coli; Fundamental and applied biological sciences. Psychology; Gene expression; Gene Expression Profiling; Hospitalization; Humans; Injuries; Investigative techniques, diagnostic techniques (general aspects); Leukocytes; Leukocytes - chemistry; Leukocytes - cytology; Medical personnel; Medical sciences; Microfluidic Analytical Techniques; Nucleic acids; Oligonucleotide Array Sequence Analysis; Patients; RNA - isolation & purification; Wounds, Nonpenetrating - pathology; Human; Leukocyte; Critically ill; Clinical biology; Isolation; Biochemistry; Hospitalization; Molecular biology; Gene expression
• ISSN: 0009-9147; 1530-8561; 1530-8561
• DOI: 10.1373/clinchem.2007.099150

Background: Microarray technology is becoming a powerful tool for diagnostic, therapeutic, and prognostic applications. There is at present no consensus regarding the optimal technique to isolate nucleic acids from blood leukocyte populations for subsequent expression analyses. Current collection and processing techniques pose significant challenges in the clinical setting. Here, we report the clinical validation of a novel microfluidic leukocyte nucleic acid isolation technique for gene expression analysis from critically ill, hospitalized patients that can be readily used on small volumes of blood. Methods: We processed whole blood from hospitalized patients after burn injury and severe blunt trauma according to the microfluidic and standard macroscale leukocyte isolation protocol. Side-by-side comparison of RNA quantity, quality, and genome-wide expression patterns was used to clinically validate the microfluidic technique. Results: When the microfluidic protocol was used for processing, sufficient amounts of total RNA were obtained for genome-wide expression analysis from 0.5 mL whole blood. We found that the leukocyte expression patterns from samples processed using the 2 protocols were concordant, and there was less variability introduced as a result of harvesting method than there existed between individuals. Conclusions: The novel microfluidic approach achieves leukocyte isolation in <25 min, and the quality of nucleic acids and genome expression analysis is equivalent to or surpasses that obtained from macroscale approaches. Microfluidics can significantly improve the isolation of blood leukocytes for genomic analyses in the clinical setting.