The application of a photon-counting camera in very sensitive, bioluminescence-enhanced detection systems for protein blotting. Ultrasensitive detection systems for protein blotting and DNA hybridization, II.
A relatively simple, very sensitive bioluminescence-enhanced detection system for protein blots was described recently. This method utilizes antibodies conjugated with alkaline phosphatase. Alkaline phosphatase releases D-luciferin (Photinus pyralis) from D-luciferin-O-phosphate. Liberated D-luciferin reacts with luciferase, ATP and oxygen with light emission. The light produced is measured with a very sensitive photon counting camera (Argus 100), permitting visualization and localization of the alkaline phosphatase-conjugated antibodies on nitrocellulose sheets. Under non-optimized conditions the limit of detection is at present 5 to 500 fg of protein (rabbit immunoglobulin G), corresponding to 30 to 3 amol. The method is therefore 10(5) times more sensitive than other used at present.
Hauber
R
R
Abteilung für Klinische Chemie, Chirurgischen Klinik Innenstadt, Universität München.
Miska
W
W
Schleinkofer
L
L
Geiger
R
R
D004247:DNA; D004586:Electrophoresis; D049454:Firefly Luciferin; D009693:Nucleic Acid Hybridization; D010783:Photometry; D011506:Proteins / Q000032:analysis*
D011506:Proteins; D049454:Firefly Luciferin; D004247:DNA