Applied Biosystems

About: Applied Biosystems is a based out in . It is known for research contribution in the topics: Mass spectrometry & Nucleic acid. The organization has 1521 authors who have published 1579 publications receiving 285423 citations.

The use of fluorescence detection and internal lane standards to size PCR products automatically.

Abstract: We have developed chemical procedures, optical and electrophoretic instrumentation and computer software automate the analysis of polymerase chain reaction (PCR) products. DNA molecules labeled with up to four different fluorescent dyes are analyzed within a single electrophoresis gel lane. A size calibration curve is created for each electrophoresis lane from the electrophoretogram of uniquely labeled DNA fragments belonging to an internal lane standard that co-electrophoreses with the PCR products. The unknown molecular lengths of PCR products are automatically calculated from the calibration curve. Data from control experiments with DNA segments of known molecular length demonstrate the accuracy and precision of such sizing. This system has been applied to the analysis of PCR products for research in the areas of human identification, genetic mapping and genetic disease.

Methods and compositions for analyzing nucleic acids

Abstract: Methods and compositions for probe amplification to detect, identify, quantitate, and/or analyze a targeted nucleic acid sequence. After hybridization between a probe and the targeted nucleic acid, the probe is modified to distinguish hybridized probe from unhybridized probe. Thereafter, the probe is amplified. Moreover, in specific embodiments, the present invention involves a chimeric probe that is particularly effective when the targeted nucleic acid sequence is short and/or has a relatively low concentration, such as with an miRNA molecule.

Duchenne/Becker muscular dystrophy carrier detection using quantitative PCR and fluorescence-based strategies.

Abstract: Dystrophin gene deletions account for up to 68% of all Duchenne (DMD) and Becker (BMD) muscular dystrophy mutations. In affected males, these deletions can be detected easily using multiplex PCR tests which monitor for exon presence. In addition, quantitative dosage screening can discriminate female carriers. We previously analyzed multiplex PCR products by gel electrophoresis and quantitation of fluorescently labeled primers with the Gene Scanner™ in order to test carrier status. These multiplex PCR protocols detect DMD gene deletions adequately, but require up to 18 pairs of fluorochrome-labeled primers. We previously described two alternative fluorescent labeling strategies, each with approximately 1,000-fold greater sensitivity than ethidium bromide staining, which can be used to quantify the products of multiplex PCR. The first method uses the DNA intercalating thiazole orange dye TOTO-1 to stain PCR products after 20 cycles. In the second method, fluorescein-12,2′-dUTP is incorporated into products during PCR as a fluorescent tag for subsequent quantitative dosage studies. Both methods label all multiplexed exons including the 506 bp exon 48 fragment that is difficult to detect and quantify by standard ethidium bromide staining. Using this approach, we determined DMD/BMD carrier status in 24 unrelated families using a fluorescent fragment analyzer. Analysis of fluorochrome-labeled PCR products facilitates quantitative multiplex PCR for gene-dosage analysis. © 1993 Wiley-Liss, Inc.

Fibulin-4 is a target of autoimmunity predominantly in patients with osteoarthritis.

Abstract: Autoimmunity to chondrocyte-producing proteins has been reported in patients with osteoarthritis (OA) as well as in those with rheumatoid arthritis (RA). To answer whether or not OA-specific autoimmunity exist, we performed screening of chondrocyte-producing autoantigens by two-dimensional electrophoresis and Western blotting with each of 20 OA and 20 RA serum samples. We identified an apparently OA-specific autoantigen spot with a molecular mass of 52 kDa and a Isoelectric point of 4.1 as fibulin-4 by mass fingerprinting. By preparing recombinant proteins of fibulin-4, we determined prevalence of the autoantibodies to fibulin-4 in 92 patients with OA, 67 patients with RA, 40 patients with systemic lupus erythematosus, and 43 patients with systemic scleroderma. As a result, the IgG type anti-fibulin-4 autoantibodies were detected in 23.9% of sera from patients with OA, in 8.9% of sera from patients with RA, in 2.5% of sera from patients with systemic lupus erythematosus, and in 9.3% of sera from patients with systemic scleroderma. Furthermore, we immunized DBA/1J, ICR, BALB/c, and C57BL/6 mice with the recombinant fibulin-4 proteins to investigate arthritogenecity of fibulin-4. As a result, mild synovitis was detected in all of the four strains. In addition, we demonstrated expression of fibulin-4 in chondrocytes at both mRNA and protein levels in vivo and in vitro by RT-PCR, Western blotting, and immunohistochemistry. Taken together, fibulin-4, expressed in chondrocytes and recognized as an autoantigen mainly in OA rather than in RA, may play pathogenic roles in OA.