Cell-free Apoptosis Assay with Xenopus Extracts

A cell-free apoptosis system was employed [44 (link),45 (link)]. Cytosolic fractions from xenopus oocytes extracts were prepared and graciously supplied by Dr. Leta Nutt at St. Jude Hospital [44 (link),45 (link)]. Purified C3 was pre-incubated with either purified cytochrome c (Sigma, MO, USA) or a purified cytosolic fraction for 1 h, followed by either addition of purified cytosol from xenopus or purified cytochrome c, and incubation for one hour. Apoptosis activities were detected using substrate from the Caspase-3 Glo Apoptosis kit (Promega, WI, USA).

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Fang Z., Lee H., Liu J., Wong K.A., Brown L.M., Li X., Xiaoli A.M., Yang F, & Zhang M. (2023). Complement C3 Reduces Apoptosis via Interaction with the Intrinsic Apoptotic Pathway. Cells, 12(18), 2282.

Publication 2023

purified cytochrome c cytochrome c

Apoptosis Caspase 3 Cell free system Cytochrome c Cytosol Oocytes Promega Xenopus

Corresponding Organization : SUNY Downstate Health Sciences University

Other organizations : Columbia University, Albert Einstein College of Medicine

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Variable analysis

independent variables

Purified C3 pre-incubated with either purified cytochrome c or a purified cytosolic fraction for 1 h

dependent variables

Apoptosis activities detected using substrate from the Caspase-3 Glo Apoptosis kit

control variables

Cytosolic fractions from xenopus oocytes extracts
Purified cytochrome c
Purified cytosol from xenopus

controls

Positive control: Purified cytochrome c
Negative control: Purified cytosolic fraction

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