Joshua A Bornhorst

Publications (25) View all

• Article: α1-Antitrypsin Phenotypes and Associated Serum Protein Concentrations in a Large Clinical Population.

  Joshua A Bornhorst, Dina N Greene, Edward R Ashwood, David G Grenache
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  ABSTRACT: α1-Antitrypsin (AAT) deficiency variants reduce the concentration of serum AAT protease inhibitor and can lead to the development of pulmonary and hepatic disease. Relative frequencies of rare AAT variant phenotypes (non-M, Z, and S) and associated serum concentrations in the clinical population have not been thoroughly described. Protein phenotypes were determined by isoelectric focusing electrophoresis for 72,229 consecutive samples. Phenotype frequencies, median serum concentrations, and central 95% concentration intervals were determined for observed phenotypes. Concurrent AAT phenotype and concentration data were used to evaluate the efficacy of using serum AAT concentration alone to detect AAT deficiency. Age, race, and sex had only slight effects on the median 95% serum protein concentration intervals of the 58,087 PiMM (wild type) phenotype specimens. Positive predictive values were calculated for the detection of potential deficiency phenotypes at different serum cutoff concentrations, aiding potential screening effort design. For example, the PiZZ deficiency phenotype (n = 814) could be detected at 99.5% sensitivity and 96.5% specificity using a cutoff of ≤ 85 mg/dL. However, at-risk specimens with two putative deleterious variants (Z, S, I, F, P, T, and Null variants) were detected with only 85.9% sensitivity at this cutoff (n = 1,661). Rare phenotype variants were observed in 2.5% of samples. This analysis provides novel information on serum AAT concentrations associated with different AAT phenotypes and provides insight into the severity of depression of AAT concentration in the presence of rare deficiency variants. Additionally, it allows for evaluation of efficacy of testing algorithms incorporating AAT serum concentration determination.
  Chest 04/2013; 143(4):1000-8. · 5.25 Impact Factor
• Article: 3-epi-25 hydroxyvitamin D concentrations are not correlated with age in a cohort of infants and adults.

  Frederick G Strathmann, Katerina Sadilkova, Thomas J Laha, Susan E LeSourd, Joshua A Bornhorst, Andrew N Hoofnagle, Rhona Jack
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  ABSTRACT: The implementation of mass spectrometry to measure serum 25-hydroxyvitamin D [25(OH)D] concentrations has led to concerns regarding the measurement and reporting of the C3-epimer of 25-hydroxyvitamin D(3) [3-epi-25(OH)D(3)], for which there is a near-total lack of data regarding its clinical significance. We developed a chromatographic method to resolve (>90%) 3-epi-25(OH)D(3) from 25(OH)D(3) using a pentafluorophenyl propyl chromatographic column. Using LC-MS/MS, we determined the serum concentrations of 25(OH)D(3) and 3-epi-25(OH)D(3) in 626 patients aged 3 days to 94 years undergoing routine vitamin D testing. Comparison between DiaSorin RIA and the new LC-MS/MS method for total 25(OH)D had acceptable agreement. Our data indicate an increase in 25(OH)D(3) rather than a reduction in epimer concentration. An average of 3.3 ng/ml of 3-epi-25(OH)D(3) was detected in adolescents and adults. Inclusion of 3-epi-25(OH)D(3) in the total 25(OH)D(3) concentration resulted in 9% (<1 year) and 3% (1 to 94 years) potential misclassification of patients as vitamin D sufficient. The new LC-MS/MS method is capable of chromatographically separating 25(OH)D(3) and 3-epi-25(OH)D(3). It was used to confirm that the contribution of 3-epi-25OHD(3) to total 25OHD(3) concentrations decreases with age in infants and is detectable in adults.
  Clinica chimica acta; international journal of clinical chemistry 01/2012; 413(1-2):203-6. · 2.54 Impact Factor
• Article: Unexplained hemolytic anemia with multiorgan failure.

  Girindra Raval, Joel E Straughen, Gwendolyn A McMillin, Joshua A Bornhorst
  Clinical Chemistry 11/2011; 57(11):1485-8. · 7.91 Impact Factor
• Article: Misclassification of an apparent alpha 1-antitrypsin "Z" deficiency variant by melting analysis.

  Dina N Greene, Melinda Procter, David G Grenache, Elaine Lyon, Joshua A Bornhorst, Rong Mao
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  ABSTRACT: Alpha-1 antitrypsin (AAT) is a protease inhibitor that protects the lungs from degradation by neutrophil elastase. AAT deficiency is associated with both lung and liver diseases. AAT deficiency is diagnosed using a combination of genetic and biochemical tests. Here we demonstrate how polymorphisms in the AAT gene can lead to genotype/phenotype discrepancies in common AAT assays. Total AAT was measured using an immunoturbidimetric assay. AAT phenotype was determined using isoelectric focusing. Genotypic identification of Z and S AAT alleles was performed by melt curve analysis using a LightCycler. Genotype/phenotype discrepancy was amended using gene sequencing. Genotype and phenotype analysis produced conflicting results as a consequence of a polymorphism located in the probe designed to detect the Z allele. Sequencing revealed that the polymorphism had previously been reported as a rare P allele. The probe that caused the discrepancy was designed to match the WT sequence. A new probe was designed to specifically detect the Z allele, eliminating the possibility of future discordance. Laboratories utilizing melt-curve analysis to diagnose patients should be aware of the potential for false positive results caused by polymorphisms located in the binding region of the genotyping probes. Alternatively, the probes should be designed to be specific to the mutation, rather than to the WT sequence.
  Clinica chimica acta; international journal of clinical chemistry 07/2011; 412(15-16):1454-6. · 2.54 Impact Factor
• Article: Immunoassay for quantifying squamous cell carcinoma antigen in serum.

  J Alan Erickson, Jun Lu, Jeffery J Smith, Joshua A Bornhorst, David G Grenache, Edward R Ashwood
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  ABSTRACT: Although the benefits of quantifying serum squamous cell carcinoma antigen (SCCa) have been reported, SCCa reagents were no longer available in the US by the late 1990s. Because SCCa quantification still has demonstrated clinical utility, we developed and validated a microtiter plate-based ELISA for measuring SCCa in serum. We coated microtiter strips overnight with capture anti-SCCa monoclonal antibody, washed the wells, added blocking buffer, and lyophilized the strips. For detection, we used a biotinylated anti-SCCa detection antibody, streptavidin/horseradish peroxidase conjugate, and tetramethylbenzidine/H(2)O(2) substrate. A novel blocking reagent against human antimouse antibodies (HAMA) was evaluated. A reference interval was established with sera from healthy individuals and was confirmed in smokers. The assay was linear to 40 microg/L SCCa (slope, 1.00; y intercept, 0.695; R(2), 0.996) with a detection limit of 0.3 microg/L. The intraassay imprecision results [mean (CV)] were 2.5 microg/L (3.4%), 18.0 microg/L (3.0%), and 30.7 microg/L (2.4%); interassay imprecision results were 2.0 microg/L (9.9%), 20.0 microg/L (7.6%), and 36.3 microg/L (3.5%). A correlation analysis against an established automated assay generated a slope of 0.976 and a y intercept of -0.193 microg/L (r(2) = 0.916). An upper reference limit of 2.1 microg/L SCCa was established at 95% confidence level, with no difference observed in smokers. No correlation between SCCa concentration and age was observed (r(2) = 0.0003). At a blocking reagent concentration of 5 mg/L, HAMA interference was eliminated in 3 samples known to produce falsely increased SCCa results. This SCCa ELISA demonstrates acceptable performance characteristics for quantifying serum SCCa and is effective in eliminating HAMA interference.
  Clinical Chemistry 09/2010; 56(9):1496-9. · 7.91 Impact Factor