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US Food and Drug Administration (FDA) as an antidote for cyanide and nitroprusside toxicity and for calciphylaxis

Biochemical Parameters

Urine samples from all groups were collected using metabolic cages for 24h and analyzed in triplicates for the levels of urea, creatinine and calcium using respective diagnostic kits from Agappe diagnostics Ltd (India). Whole blood was collected from the retro-orbital sinus on the day of necropsy, centrifuged at 10.000×g for 10 min. and serum chemistry analysis was performed in triplicates for calcium, creatinine, urea, ALP using the respective diagnostic kits purchased from Agappe diagnostics Ltd. & Span diagnostics Ltd. (India).

Antioxidant assays

After necropsy left kidney was cut into four equal sections. Each section was weighed separately, crushed and homogenized in 3mL ice cold Tris buffer (pH=7.4) for performing various assays. Total protein content was measured by Lowry et al., (1956) and used for further calculation. The remaining sample was used for the estimation of various antioxidant levels in kidney homogenate such as TBARS, SOD, GPX and catalase by previously described standard methods while the level of ALP was measured using commercial kit.

Histopathology

After 21 days of treatment, rats were euthanized by carbon dioxide inhalation followed by cervical dislocation. Immediate laparotomy was performed to collect both the kidneys. Isolated kidneys were cleaned off the extraneous tissue, weighed and rinsed with ice-cold normal saline. A section from both kidneys was fixed with 10% v/v neutral formalin and processed through graded alcohol series and xylene, embedded in paraffin, sectioned at 5μm, and stained with hematoxylin and eosin for histopathological examination under a light microscope. Three kidney tissues per group were analyzed for nephropathy, obstruction and stone deposition.