Application

This kit can be used for detection of endogenous carbon monoxide (CO) content in serum and plasma samples.

Detection significance

Carbon monoxide (CO) is a colorless, odorless, and tasteless gas that is slightly less dense than air. Carbon monoxide (CO) in tissues and cells can originate from inhalation of CO or endogenously. Endogenous production, carboxyhemoglobin (COHb) formation, and exhaled CO levels are influenced by physiological factors, including disease. It is suggested that endogenous CO production can be used as a biomarker for oxidative and inflammatory processes. Also, endogenous CO can contribute to increased body burden of CO, which may both disrupt normal CO signaling cascades and increase the risk of CO toxicity.

Detection principle

According to the difference of absorption spectrum between carboxyhemoglobin (HbCO) and reductive hemoglobin (Hb), two wavelengths with the biggest difference in HbCO absorbance and zero in HB absorbance were selected, and the absorbance difference (△OD) of samples at these two wavelengths was determined by dual wavelength spectrophotometry. The content of endogenous carbon monoxide is calculated according to the percent concentration of HbCO through standard curve.

Experimental instrument

Test tube, Micropipettor, Vortex mixer, Water bath, Spectrophotometer (568 nm and 581 nm)

Operating steps

1. Detection of hemoglobin (Hb) in serum(plasma) sample:

Take 0.05 mL of sample (serum or plasma), add 2.5 mL of Reagent 5 application solution, mix fully and stand for 5 min, set to zero with double distilled water and measure the OD values of each tube at 540 nm with 1 cm cuvette.

2. Pretreatment of sample:

(1) Preparation of hemolysis blood: take 0.01 mL of anticoagulant whole blood (Don't centrifuge), add 1.2 mL of Reagent 1, mix thoroughly and stand for 5 min to prepare hemolysis blood.

(2) Preparation of tested sample: take 0. 1 mL of prepared hemolysis blood, add 0.1 mL of plasma (or serum) and mix fully.

(3) Preparation of control sample: take 0. 1 mL of prepared hemolysis blood, add 0.1 mL of double distilled water and mix fully.

3. Detection of HbCO in serum(plasma) samples:

Mix fully and stand for 10 min, set to zero with Reagent 2 and measure the OD values of each tube with 1 cm cuvette at 568 nm and 581 nm respectively. Then calculate the absolute △OD value:

△OD= (OD568 of sample – OD581 of sample) - (OD568 of control – OD581 of control)