Analysis of transforming growth factor-beta 1 (TGF-beta1) expression in human monocytes infected with Mycobacterium avium at a single cell level by ELISPOT assay.

Transforming growth factor beta 1 (TGF-beta1) has been implicated in the pathogenesis of a number of diseases including infection with intracellular pathogens such as Mycobacterium avium complex (MAC). In this study, we developed an ELISPOT assay for measurement of active TGF-beta1 produced by peripheral blood mononuclear cells (PBMC) from healthy individuals in response to LPS or MAC. The frequency of TGF-beta1 producing cells was significantly (p<0.04) higher in response to LPS (10 microg/ml) as compared to unstimulated cells (n=4). Moreover, the frequency of TGF-beta1 producing cells was threefold higher in monocyte (MN)-enriched cell population than those in PBMC indicating that the source of TGF-beta1 producing cells in PBMC was MN. In addition, the frequency of TGF-beta1 producing cells in response to MAC (10:1, cfu:MN) was significantly higher (p<0.03) than unstimulated cells. However, the frequency of TGF-beta1 producing cells in response to MAC (10:1) was eight to ninefold lower than that by LPS (10 microg/ml). Moreover, there was a correlation between the level of total TGF-beta1 in 24-h culture supernatants and the number of TGF-beta1 producing cells upon MAC stimulation. TGF-beta1 ELISPOT-assay may be a sensitive and a powerful tool for detection of TGF-beta1 producing cells, and may be helpful in elucidation of the nature of TGF-beta1 production at sites of diseases.     

BRAF mutation testing with a rapid,fully integrated molecular diagnostics system

Fast and accurate diagnostic systems are needed for further implementation of precision therapy of BRAF-mutant and other cancers. The novel Idylla^TM BRAF Mutation Test has high sensitivity and shorter turnaround times compared to other methods. We used Idylla to detect BRAF V600 mutations in archived formalin-fixed paraffin-embedded (FFPE) tumor samples and compared these results with those obtained using the cobas 4800 BRAF V600 Mutation Test or MiSeq deep sequencing system and with those obtained by a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory employing polymerase chain reaction–based sequencing, mass spectrometric detection, or next-generation sequencing. In one set of 60 FFPE tumor samples (15 with BRAF mutations per Idylla), the Idylla and cobas results had an agreement of 97%. Idylla detected BRAF V600 mutations in two additional samples. The Idylla and MiSeq results had 100% concordance. In a separate set of 100 FFPE tumor samples (64 with BRAF mutation per Idylla), the Idylla and CLIA-certified laboratory results demonstrated an agreement of 96% even though the tests were not performed simultaneously and different FFPE blocks had to be used for 9 cases. The Idylla^TM BRAF Mutation Test produced results quickly (sample to results time was about 90 minutes with about 2 minutes of hands on time) and the closed nature of the cartridge eliminates the risk of PCR contamination. In conclusion, our observations demonstrate that the Idylla test is rapid and has high concordance with other routinely used but more complex BRAF mutation–detecting tests.     

Killing two birds with one stone: the potential effect of cataract surgery on the incidence of primary angle-closure glaucoma in a high-risk population

Background: To estimate the proportion of cataract surgery performed at various visual acuity and lens opacity thresholds that would coincidentally treat early angle‐closure disease, and to estimate the effect of this surgery on the incidence of primary angle‐closure glaucoma. Design: Cross‐sectional, population‐based survey in Meiktila, Myanmar. Participants: Total of 2076 inhabitants, 40 years of age and over were included. Methods: Eyes with cataract‐induced visual impairment, and primary angle‐closure disease were identified. Analyses were stratified by various pinhole‐corrected visual acuity and Lens Opacity Classification System III scores thresholds. Main Outcome Measures: The dual role of cataract surgery in primary cataract treatment and primary angle‐closure glaucoma prevention was estimated. Results: Of 4153 eyes available for analysis, 261 eyes were either primary angle‐closure suspect or primary angle closure; 975 eyes had a visual acuity of <6/18 and Lens Opacity Classification System III score ≥3 on the nuclear or cortical scales. Of these, 86 eyes had either primary angle‐closure suspect or primary angle closure. If cataract surgery were performed on all 975 eyes, this would potentially prevent up to 86 cases of primary angle‐closure glaucoma in this population; 8.82% (95% confidence interval 7.12–10.78%) of the cataract surgery would address the cataract and prevent primary angle‐closure glaucoma. This would achieve a 38.46% (95% confidence interval 20.23–59.43%) relative reduction in the incidence of primary angle‐closure glaucoma in the adult population. Conclusion: In populations with a high prevalence of both visually significant cataract and angle‐closure disease, quality cataract extraction can serve a dual role of visual restoration and reducing the incidence of angle‐closure disease in the population: killing two birds with one stone.