Nanodrop 1000 uv vis spectrophotometer software

Total RNA was isolated from DCs in vitro and from oral mucosal (gingiva) tissue of the experimental groups used for in vivo studies using QIAGEN RNeasy mini kit (Qiagen, Inc., Valencia, CA, and USA). RNA purity and concentration were analyzed with Nanodrop (NanoDrop 1000 UV-VIS Spectrophotometer Software Ver.3.8.1, Thermofisher Scientific). Ratio of 260/280 of 2.0 was considered adequate for analysis and was reverse transcribed to cDNA. Amplification by PCR was performed using the High-Capacity cDNA Reverse Transcription Kit and PCR in total reaction of 20 μL. Quantitative real-time PCR was performed using TaqMan gene expression primers specific for ACE 2 (Mm00446190_m1), and Beta Actin (Mm02619580_g1)(Thermofisher Scientific). RT-PCR was run in StepOnePlus Real-Time PCR System. Relative gene expression was determined using delta-delta CT and plotted as relative fold change.

Total RNA was isolated using QIAGEN RNeasy mini kit (Qiagen, Inc., Valencia, CA, and USA). RNA purity and concentration were measured using Nanodrop (NanoDrop 1000 UV-VIS Spectrophotometer Software Ver.3.8.1, Thermofisher Scientific). Ratio of 260/280 of 2.0 was considered adequate for analysis. Reverse transcription to cDNA was performed using the High-Capacity cDNA Reverse Transcription Kit (Applied Biosystem, Thermofisher Scientific, Waltham MA, USA) in total reaction of 20 μL. Quantitative real-time PCR was performed using TaqMan fast advanced master mix (Applied Biosystem, Thermofisher Scientific, Waltham MA, USA) and Taq-Man Gene Expression assay (Applied Biosystem, Foster City, CA, USA) specific for: IL6 (Mm00446190_m1), TNF (Mm00443258_m1) and IL1B (Mm00434228_m1), internal control Glyceraldehyde-3-phosphate dehydrogenase (GAPDH Mm99999915_m1). RT-PCR was run in StepOnePlus Real-Time PCR System. Calculation of relative mRNA expression was performed using delta-delta CT and presented as relative fold-change.

Total RNA was isolated from DCs using a QIAGEN RNeasy mini kit (Qiagen, Inc., Valencia, CA, USA; Cat#: 74134). A Nanodrop instrument (NanoDrop 1000 UV-VIS Spectrophotometer Software Ver.3.8.1, Thermofisher Scientific) was used to assess RNA concentration and purity. A ratio of 260/280 of 2.0 was considered suitable for further analysis. A High-Capacity Reverse Transcription Kit (Applied Biosystem, Thermofisher Scientific, Waltham MA, USA) was used to perform reverse transcription to cDNA in a total reaction volume of 20 µL. Quantitative real-time PCR was performed using TaqMan fast advanced master mix (Applied Biosystem, Thermofisher Scientific, Waltham, MA, USA) and TaqMan Gene Expression assays (Applied Biosystem, Foster City, CA, USA) specifically for IL6 (Hs00174131_m1), IL1 (Hs000961622_m1), IL-1B (Hs01555410_m1), TNFa (Hs01113624_g1), IL23 (Hs0372324_m1), and internal control GADPH (Hs02758991_g1). RT-PCR was run in a StepOnePlus Real-Time PCR System. Calculation of relative gene expression was performed using delta-delta CT and plotted as relative fold change.