A Spectrophotometric Assay for Robust Viability Testing of Seed Batches Using 2,3,5-Triphenyl Tetrazolium Chloride: Using Hordeum vulgare L. as a Model

Abstract

A comparative analysis was carried out of published methods to assess seed viability using 2,3,5-triphenyltetrazolium chloride (TTC) based assays of seed batches. The tests were carried out on seeds of barley (Hordeum vulgare cv. Optic) as a model. We established that 10% [w/v] trichloroacetic acid (TCA)/methanol is superior to the acetone and methanol-only based methods: allowing the highest recovery of formazan and the lowest background optical density (OD) readings, across seed lots comprising different ratios of viable and dead seeds. The method allowed a linear-model to accurately capture the statistically significant relationship between the quantity of formazan that could be extracted using the method we developed and the seed temperature-response, and seed viability as a function of artificially aged seed lots. Other quality control steps are defined to help ensure the assay is robust and these are reported in a Standard Operating Procedure.

Maternal transfer of environmentally relevant polybrominated diphenyl ethers (PBDEs) produces a diabetic phenotype and disrupts glucoregulatory hormones and hepatic endocannabinoids in adult mouse female offspring

“…GDH is a key enzyme bridging amino acid-to-glucose pathways. GDH activity in crude liver homogenates was assayed using the tetrazolium salt method of Lee and Lardy 98 with modification for multi-well plates 99 . The 5% (w/v) homogenates of liver (5 mg) were prepared in 0.25 M sucrose solution.…”

“…The reaction, in all the samples, was started by the addition of sample liver homogenate and proceeded for 30 min at 37 °C. The formazan formed was measured as optical density at 545 nm then converted to concentration using a iodonitrotetrazolium formazan (TCI) standard curve fitted with a linear regression model 99 . Protein content was measured using a bicinchoninic acid assay (ThermoFisher).…”

Growth promotion and disease resistance induced in Anthurium colonized by the beneficial root endophyte Piriformospora indica

“…The root vigor was determined by TTC (2,3,5-triphenyltetrazolium chloride) methods [17]. The roots were washed with distilled water and patted dry on the filter paper.…”

Maternal Transfer of Environmentally Relevant Polybrominated Diphenyl Ethers (PBDEs) Produces a Diabetic Phenotype and Disrupts Glucoregulatory Hormones and Hepatic Endocannabinoids in Adult Mouse Female Offspring

“…GDH is a key enzyme bridging amino acid-toglucose pathways. GDH activity in crude liver homogenates was assayed using the tetrazolium salt method of Lee and Lardy 98 with modification for multi-well plates 99 exposure produced significant accumulation of PBDEs in liver, being greater in directly exposed F0 versus indirectly exposed F1 female mice. Dose-dependency was only seen in F1.…”