A multiplexed protein microarray for the simultaneous serodiagnosis of human immunodeficiency virus/hepatitis C virus infection and typing of whole blood.

All donor blood samples must be tested pretransfusion to determine the donor blood type. Standard testing protocols require that assays be performed for important bloodborne pathogens such as hepatitis C, syphilis, hepatitis B, and human immunodeficiency virus. We have demonstrated proof of the concept that a protein microarray can type whole blood and detect antibody to significant pathogens simultaneously from the same donor blood sample. The data collected demonstrate the ability of the array to accurately type blood samples while also detecting the presence of antibodies against both human immunodeficiency virus and hepatitis C virus. In conclusion, we have successfully developed a platform capable of typing human whole blood samples, while at the same time testing for the presence of antibodies specific for human immunodeficiency virus/hepatitis C virus. The major benefits of this system are its amenability to expansion with additional assays, for example, rhesus typing and syphilis and/or hepatitis B virus detection, and also the adaptability of the assay to higher-throughput analysis, currently 16 individual samples per slide, but readily expandable to a 96-well format.

Evaluation of a protein microarray method for immuno-typing erythrocytes in whole blood.

All donor blood samples must be tested pre-transfusion to determine the blood type of donor erythrocytes, based on the ABO typing system. Current methods of testing are well characterised, but require a number of processing steps prior to analysis. In addition, standard testing protocols require additional assays such as hepatitis C and HIV testing be performed separately. We describe and evaluate a protein microarray platform for ABO blood typing that has the potential to be a simple reliable high throughput method, with the added capability for the integration of other important pre-transfusion tests. Sixty seven donor blood samples were incubated on microarrays printed with multiple spotted replicates of blood type antigen specific antibodies. We utilised a hold-out cross validation approach, combined with Receiver Operator Characteristic (ROC) curves to define thresholds within which a sample could be defined as being of a particular blood type. The threshold values from the ROC curve analysis demonstrated an excellent ability to accurately separate samples based on ABO blood type. The results obtained when the thresholds from the training sets were applied to test sets were also very encouraging, with misclassified samples being present in only 2 of the training sets and a mean classification error of 4.28%. When the mean thresholds were applied to the 67 donor samples, 95.5% were correctly blood typed (64 of 67 samples). We have demonstrated the ability of our protein microarray platform to successfully and accurately type human whole blood samples. We believe that this flexible platform provides a strong basis for an integrated approach for combined blood typing and pathogen testing in human whole blood.

Cell interaction microarray for blood phenotyping.

Microarrays promise great advances in areas of diagnostic testing where there is a need to perform multiple assays in parallel. In the short term, protein microarrays have a greater potential to impact diagnostics than DNA arrays due to their potential for direct sample measurements. Here, we report an antibody microarray technique for selectively recognizing glycan and peptide motifs on the surface of red blood cells. We present results demonstrating the optimization and efficacy of the microarray approach as a highly sensitive and specific microscale multiplex assay for blood typing. We also show that our microarray can be used to screen red blood cell surface antigens using whole blood in a label-free detection mode. Finally, our results indicate this method has potential for broader applications in biochip medicine.

The novel herbicide oxaziclomefone inhibits cell expansion in maize cell cultures without affecting turgor pressure or wall acidification.

Oxaziclomefone [OAC; IUPAC name 3-(1-(3,5-dichlorophenyl)-1-methylethyl)-3,4-dihydro-6-methyl-5-phenyl-2H-1,3-oxazin-4-one] is a new herbicide that inhibits cell expansion in grass roots. Its effects on cell cultures and mode of action were unknown. In principle, cell expansion could be inhibited by a decrease in either turgor pressure or wall extensibility. Cell expansion was estimated as settled cell volume; cell division was estimated by cell counting. Membrane permeability to water was measured by a novel method involving simultaneous assay of the efflux of (3)H(2)O and [(14)C]mannitol from a 'bed' of cultured cells. Osmotic potential was measured by depression of freezing point. OAC inhibited cell expansion in cultures of maize (Zea mays), spinach (Spinacia oleracea) and rose (Rosa sp.), with an ID(50) of 5, 30 and 250 nm, respectively. In maize cultures, OAC did not affect cell division for the first 40 h. It did not affect the osmotic potential of cell sap or culture medium, nor did it impede water transport across cell membranes. It did not affect cells' ability to acidify the apoplast (medium), which may be necessary for 'acid growth'. As OAC did not diminish turgor pressure, its ability to inhibit cell expansion must depend on changes in wall extensibility. It could be a valuable tool for studies on cell expansion.

Oxaziclomefone, a new herbicide, inhibits wall expansion in maize cell-cultures without affecting polysaccharide biosynthesis, xyloglucan transglycosylation, peroxidase action or apoplastic ascorbate oxidation.

BACKGROUND AND AIMS: Oxaziclomefone (OAC), a new herbicide, inhibits cell expansion, especially in roots and cell-cultures of gramineous monocots. OAC does not affect turgor in cultured maize cells, and must therefore inhibit wall-loosening or promote wall-tightening. METHODS: The effects of OAC in living cultured maize cells on various biochemical processes thought to influence wall extension were studied. KEY RESULTS: OAC did not affect 14C-incorporation from D-[U-14C]glucose into the major sugar residues of the cell wall (cellulosic glucose, non-cellulosic glucose, arabinose, xylose, galactose, mannose or uronic acids). OAC had no effect on 14C-incorporation from trans-[U-14C]cinnamate into wall-bound ferulate or its oxidative coupling-products. OAC did not influence the secretion or in-vivo action of peroxidase or xyloglucan endotransglucosylase activities-proposed wall-tightening and -loosening activities, respectively. The herbicide did not affect the consumption of extracellular L-ascorbate, an apoplastic solute proposed to act as an antioxidant and/or to generate wall-loosening hydroxyl radicals. CONCLUSIONS: OAC decreased wall extensibility without influencing the synthesis or post-synthetic modification of major architectural wall components, or the redox environment of the apoplast. The possible value of OAC as a probe to explore aspects of primary cell wall physiology is discussed.