Efficient assessment of the utility of immortalized Fa2N-4 cells for cytochrome P450 (CYP) induction studies using multiplex quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) and substrate cassette methodologies

Xenobiotica ◽  
2008 ◽  
Vol 38 (12) ◽  
pp. 1500-1517 ◽  
Author(s):  
J. R. Kenny ◽  
L. Chen ◽  
D. F. McGinnity ◽  
K. Grime ◽  
K. M. Shakesheff ◽  
...  
2020 ◽  
Vol 40 (4) ◽  
pp. 227-233
Author(s):  
Claudia S. Wisser ◽  
André Thaler Neto ◽  
Helena B.C.R. Batista ◽  
Enio Mori ◽  
Maria E.R. Chierato ◽  
...  

ABSTRACT: Our objective was the characterization and staging of histological lesions in different anatomical sites of the central nervous system (CNS) of rabid cattle. The severity of the lesions was compared with the clinical stages of the disease, the variants of viral isolates, and with the load of virus. Thirty-one spontaneously affected rabid cattle the state of Santa Catarina underwent clinical follow-up and were eventually necropsied. CNS tissues were sampled and submitted to direct fluorescent antibody technique (DFAT), immunohistochemistry (IHC), routine histopathology with hematoxylin and eosin stain (HE), reverse transcriptase polymerase chain reaction (RT-PCR), and polymerase chain reaction in quantitative reverse transcriptase in real time (qRT-PCR). Affected cattle were allotted in four groups according to their clinical stage when euthanized: G1, euthanized while standing; G2, euthanized when in sternal recumbence; G3, euthanized when in lateral recumbence; and G4, affected cattle with natural death. In order to evaluate the degree of severity of the lesions and the presence of Negri bodies (NBs), the brain was sectioned at 9 sites. Additionally, spinal cord and trigeminal ganglion sections were examined. The intensity of the lesions was graded as either absent, mild, moderate, or marked, and the presence or absence of the NBs was noted. Histological lesions were characterized by lymphocytic and monocytic meningoencephalitis with NBs in 28 cases. In all analyzed groups, intensities of histological lesions ranging from mild to severe were observed. Brain regions with the highest inflammatory lesion intensity were the medulla at the level of obex, followed by the colliculus and thalamus. NBs were observed in a higher percentage in the cerebellum, followed by medulla at the obex level, striatum complex, and frontal telencephalon. The duration of the clinical course of the disease did not influence the intensity of the inflammatory lesion, but it did influence the presence of NBs, with a higher percentage of these inclusions in cattle that died naturally than in euthanized cattle. All isolated rhabdovirus included in this study were genetically compatible with samples from hematophagous bats Desmodus rotundus. The evaluation by qRT-PCR did not demonstrate a correlation between lesion intensity and the amount of virus.


Author(s):  
Eelke Brandsma ◽  
Han J M P Verhagen ◽  
Thijs J W van de Laar ◽  
Eric C J Claas ◽  
Marion Cornelissen ◽  
...  

Abstract Background Recent advances in CRISPR-based diagnostics suggest that DETECTR, a combination of reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) and subsequent Cas12 bystander nuclease activation by amplicon-targeting ribonucleoprotein complexes, could be a faster and cheaper alternative to quantitative reverse-transcription polymerase chain reaction (qRT-PCR) without sacrificing sensitivity and/or specificity. Methods In this study, we compare DETECTR with qRT-PCR to diagnose coronavirus disease 2019 on 378 patient samples. Patient sample dilution assays suggest a higher analytical sensitivity of DETECTR compared with qRT-PCR; however, this was not confirmed in this large patient cohort, where we report 95% reproducibility between the 2 tests. Results These data showed that both techniques are equally sensitive in detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) providing additional value of DETECTR to the currently used qRT-PCR platforms. For DETECTR, different guide ribonucleic acids can be used simultaneously to obviate negative results due to mutations in N-gene. Lateral flow strips, suitable as a point-of-care test, showed a 100% correlation to the high-throughput DETECTR assay. More importantly, DETECTR was 100% specific for SARS-CoV-2 relative to other human coronaviruses. Conclusions Because there is no need for specialized equipment, DETECTR could be rapidly implemented as a complementary technically independent approach to qRT-PCR thereby increasing the testing capacity of medical microbiological laboratories and relieving the existent PCR platforms for routine non-SARS-CoV-2 diagnostic testing.


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