Rapid spectrophotometric method for quantitation of cytochrome c release from isolated mitochondria or permeabilized cells revisited

Abstract

This paper recalls the earlier work by Keilin, Margoliash and others at the beginning of the 20th century and shows how their results can be used for the rapid solution of new problems of modern science. It describes a rapid and simple spectrophotometric method for quantitative determination of cytochrome c release from isolated mitochondria or permeabilized cells induced by proapoptotic proteins. For this, the Soret (gamma) peak at 414 nm in the spectrum of cytochrome c is used. The results of spectrophotometric assay of cytochrome c release are in accord with those of oxygraphic determination of cytochrome c-dependent respiration of isolated mitochondria and permeabilized cardiomyocytes.

Ganoderic acid T from Ganoderma lucidum mycelia induces mitochondria mediated apoptosis in lung cancer cells

“…After induction treatment, cells were treated as reported (Appaix et al, 2000;Elliott et al, 2003). Cells (2 × 10 6 ) were rinsed three times with PBS, scraped from the dish and then lysed and centrifuged cells for separation of mitochondrion (4°C, 25,000 g, 30 min).…”

Arachidonic Acid Causes Cytochrome c Release from Heart Mitochondria

“…2B). It has to be noted that mitochondria subjected to hypotonic rupture in saline medium (29) released almost the same amount of cytochrome c (not shown). Quite consistently with the release values in the supernatant, it was found that around 46% (Ϯ4%, n ϭ 8) of the total cytochrome c content remained in the pellet (42% in the experiment of Fig.…”

Cytotoxic effect of a mannogalactoglucan extracted from Agaricus bisporus on HepG2 cells

“…Cytochrome c release was monitored spectrophotometrically by the variation in absorbance at 414 nm, according to Appaix et al (2000). HepG2 cells (1x10 6 /plate) were seeded in a 60 mm plate and incubated with RK2-Ab for 24 h. After incubation, cells were harvested using a trypsin solution and centrifuged at 1200 rpm for 10 min at 4 °C.…”