Detection of chemiluminescence efficiency of acridinium ester

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We know that chemiluminescence immunoassay is a relatively advanced detection method in the field of in vitro diagnostics, and acridine ester is a reagent with high detection sensitivity and high quantum yield among chemiluminescence reagents, and its luminescence reaction does not require enzyme catalysis, so acridine How to measure the luminous efficiency of pyridinyl ester?

Chemiluminescence efficiency detection of acridinium ester:

Use a photometer to measure the number of light quanta per second after the acridine ester is excited, and measure the luminous intensity of different concentration and volume respectively to obtain its luminous efficiency.

1. First, prepare the reagents required for the chemiluminescence experiment, including: dimethylformamide DMF, purity> 99%; hydrogen peroxide, nitric acid, sodium hydroxide, analytical purity; cetyl trimethyl ammonium chloride ( Ctac); Tween-20, TritonX-100; 96-well plate; anti-human TSH monoclonal antibody.

2. Take a certain amount of DMAE-NHS solution in a microwell plate, use a black opaque microwell plate for a 96 microwell plate, add a luminescence starting reagent, and measure the luminescence count over time on a luminometer. A similar method is used to measure NSP-DMAE-NHS or other types of acridinium esters.

3. By changing the concentration and volume of each component (H2O2, HNO3 and NaOH) in the luminescence starting reagent, respectively measuring the luminous intensity corresponding to different concentration and volume, the influence of the luminescence starting reagent and surfactant on the luminescence intensity of DMAE-NHS can be analyzed . Add the surfactants Ctac, Tween-20, TritonX-100 to the luminescence starting reagent, and measure the luminescence intensity.

Experiments of Reagents in Chemiluminescence of Acridinium Ester

In the acridinium ester luminescence experiment, hydrogen peroxide is the oxidant that stimulates chemiluminescence, sodium hydroxide makes the pH of the system in an alkaline environment, and the addition of hydrochloric acid or nitric acid to the stimulator has a certain promoting effect; in addition, the surfactants Ctac and Tween are added. -20. TritonX-100 can also enhance chemiluminescence, but Ctac will increase the background, so it is not suitable as an acridine ester DMAE-NHS chemiluminescence enhancer.

The chemiluminescence efficiency of acridinium ester is very high, but it is very different from the chemiluminescence of luminol. Luminol is a glow type with a long luminescence duration, and acridine ester is a flash type with a luminescence time of only 2-4 Second, it needs to be detected immediately with a photometer. In addition to DMAE-NHS, there are 5 kinds of acridinium esters developed by Desheng, which can be used to label different kinds of proteins, antibodies and nucleic acids.