Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2012 Jan;64(1):9-14.
doi: 10.1007/s10616-011-9395-0. Epub 2011 Sep 24.

An improved non-enzymatic "DNA ladder assay" for more sensitive and early detection of apoptosis

Shubhankar Suman  1 , Akshay Pandey, Sudhir Chandna
Affiliations

An improved non-enzymatic "DNA ladder assay" for more sensitive and early detection of apoptosis

Shubhankar Suman et al. Cytotechnology. 2012 Jan.

Abstract

Conventional DNA ladder assay has certain shortcomings such as loss of DNA fragments during sample processing, involvement of multiple steps and requirement of expensive reagents. The present study demonstrates a rapid, easy-to-perform cost-effective method for detection of apoptotic DNA fragments with considerable improvement in the sensitivity by avoiding loss of DNA fragments. It involves a few minutes of procedure involving direct lysis of cells with dimethyl sulphoxide (DMSO), brief vortexing, addition of 2% SDS-TE buffer, and a single step of centrifugation. This cost- and time-efficient method reduces the assay time considerably and can be used for a large number of samples with excellent sensitivity.

PubMed Disclaimer

Figures

Fig. 1
Fig. 1
Morphological analysis of stress-induced apoptotic body formation in Sf9 cells. Cells were treated with known apoptogenic agents; Actinomycin D (0.5 μg/mL), Etoposide (100 μM) and γ-radiation (2,000 Gy). Images were acquired with an Axiovert 200 Zeiss inverted microscope (Carl Zeiss, Germany) using Nomarski-DIC mode and Axiovision (version 4.0) software
Fig. 2
Fig. 2
DNA ladders of treated Sf9 cells obtained using the conventional phenol–chloroform method. a 3 × 106 cells were used and 40 μL of final sample was loaded in all lanes. Electrophoresis was performed on 2% agarose gel. b Densitometric analysis of ladder lanes using Image-J software, with distinct peaks representing a single band on the gel, ranging from high to low molecular weight
Fig. 3
Fig. 3
DNA ladders of treated Sf9 cells obtained using the new DMSO-SDS-TE method. a Equal volume (40 μL) of lysate was loaded in all lanes. Electrophoresis was performed on 2% agarose gel. Relative abundance of small DNA fragments is evident in the new assay compared to the conventional method. b Densitometric analysis of ladder lanes using Image-J software, with distinct peaks representing a single band on the gel, ranging from high to low molecular weight
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Arakawa T, Kita Y, Timasheff SN. Protein precipitation and denaturation by dimethyl sulfoxide. Biophys Chem. 2007;131:62–70. doi: 10.1016/j.bpc.2007.09.004. - DOI - PubMed
    1. Basnakian AG, James SJ. A rapid and sensitive assay for detection of DNA fragmentation during early phases of apoptosis. Nucleic Acids Res. 1994;22:2714–2715. doi: 10.1093/nar/22.13.2714. - DOI - PMC - PubMed
    1. Chandna S. Single-cell gel electrophoresis assay monitors precise kinetics of DNA fragmentation induced during programmed cell death. Cytometry Part A. 2004;61A:127–133. doi: 10.1002/cyto.a.20071. - DOI - PubMed
    1. Chandna S, Dwarakanath BS, Seth RK, Khaitan D, Adhikari JS, Jain V. Radiation responses of Sf9, a highly radioresistant lepidopteran insect cell line. Int J Radiat Biol. 2004;80:301–315. doi: 10.1080/09553000410001679794. - DOI - PubMed
    1. Compton MM. A biochemical hallmark of apoptosis: intranucleosomal degradation of the genome. Cancer Metast Rev. 1992;11:105–119. doi: 10.1007/BF00048058. - DOI - PubMed

LinkOut - more resources