[Frontiers in Bioscience 7, c1-12, January 1, 2002]
NITROCELLULOSE: A TRIED AND TRUE POLYMER FINDS UTILITY AS A POST-GENOMIC SUBSTRATE
John L. Tonkinson and Brett A. Stillman
Research and Development, Schleicher & Schuell BioScience, Inc., 10 Optical Ave, Keene, NH, 03431
Figure 1. Structure of monomer unit of (A.) Cellulose and (B.) Nitrocellulose. The nitrocellulose unit shows the tri-substituted congener. X = 500 - 2,500.
Figure 2. S.E.M. images of various pore size nitrocellulose membranes. (A.) Average pore size = >5micrometers; (B.) Average pore size = 1micrometer; (C.) Average pore size = 0.2micrometers; (D.) Average pore size = <0.1micrometer.
Figure 3. Schematic representation of hydrophobicity of nitrocellulose membranes. The nitrocellulose polymer is inherently hydrophobic. Aqueous solutions of biomolecules cannot penetrate a membrane that is pure nitrocellulose. If a surfactant is added during polymerization, then the hydrophobicity decreases, allowing aqueous solutions to penetrate.
Figure 4. Comparison of 2-dimensional and 3-dimensional surfaces for arraying applications. (A.) A 2-dimensional activated surface, where R could be NH2, COH, COOH, or other reactive moiety. (B.) A 3-dimensional microporous structure on a glass surface.
Figure 5. Comparison of protein microarray on nitrocellulose-coated glass (A.) vs. aldehyde-derivatized glass (B.). Cell lysates were made from CEM lymphocytic leukemia cells and then arrayed in serial dilution on the 2 surfaces. After fixation and blocking, arrays were probed using and anti-PCNA antibody followed by a Cy5 labeled secondary antibody. Slides were imaged in a ScanArray 4000 (Packard Instruments, Inc.). The Nitrocellulose slide was imaged at 25% laser power and 60% PMT voltage. The aldehyde slide was imaged at 80% laser power and 80% PMT voltage.
Figure 6. Comparison of linearity and sensitivity for sandwich assays for TNFa. Top: Standard curve from traditional colorimetric ELISA performed in a microtiter plate. Bottom: Standard curve from fluorescent sandwich assay performed on nitrocellulose thin film.