A convenient one-step extraction of cellular ATP using boiling water for the luciferin-luciferase assay of ATP

Author(s): Yang NC, Ho WM, Chen YH, Hu ML


Cellular ATP is commonly determined as production of bioluminescence using a luciferin-luciferase reaction system. Before the measurement of bioluminescence, cellular ATP must first be extracted. Two commonly used extraction methods are: () Tris-borate buffer (pH 9.2) coupled with a heating process (to inactivate ATPase) and () perchloric acid followed by neutralization. However, we found that both Tris-borate buffer and perchloric acid interfered with the luciferin-luciferase system. Here, we report a convenient single-step boiling deionized water (DW) method for extracting cellular ATP to replace perchloric acid and Tris-borate buffer. We showed that the boiling DW method did not interfere with the bioluminescence and was effective in inhibiting ATPase. This improved method required no neutralization and dilution and thus was more convenient than the perchloric acid method. Unlike the Tris-borate/heating procedure, our method did not require a separate heating step because boiling DW effectively inhibited ATPase and thus accomplished the two missions in one step for both suspended and attached cells. The improved method was precise for both suspended cells and attached cells, when cell numbers were between 10(3) and 10(6). The method also was more sensitive than other methods because it required much fewer cells (10(4) to 10(5)) than other methods for ATP determination. Thus, this one-step method is suitable for routine assay of cellular ATP for both suspended and attached cells.

Similar Articles

Molluscidal properties of some species of Annona

Author(s): Dos Santos AF, Sant’ana AE

Pure annonin and a process for the preparation thereof, US Patent 4689232

Author(s): Moeschler HF, Pfluger W, Wendisch D

Control of pests with annonaceous acetogenins, US Patent 4721727

Author(s): Mikolajczak KJ, McLaughlin JL, Rupprecht JK

Control of pests with annonaceous acetogenins, US Patent 4721727

Author(s): Mikolajczak KJ, McLaughlin JL, Rupprecht JK

Acetogenins from Annona glabra seeds, Phytochemistry 47: 811-816

Author(s): Teresa G, Raul A, Jose RT, Amparo MB, Carmen ZMP

Comparative efficacy of Annona squamosa Linn

Author(s): George S, Vincent S

Selective action of acetogenin mitochondrial complex I inhibitors

Author(s): González-Coloma A, Guadaño A, de Inés C, Martínez-Díaz R, Cortes D

Insecticidal and mutagenic evaluation of two annonaceous acetogenins

Author(s): Guadaño A, Gutiérrez C, de La Peña E, Cortes D, González-Coloma A

Medical Insects and Arachnids

Author(s): Crosskey RW, Lane RP

Protein measurement with the Folin phenol reagent

Author(s): Lowry OH, Rosebrough NJ, Farr AL, Randall RJ

Acetogenins from Annonaceae, inhibitors of mitochondrial complex I

Author(s): Zafra-Polo MC, González MC, Estornell E, Sahpaz S, Cortes D

Flics and disease

Author(s): Greenberg B