Note: Take two or three different samples for prediction before the test.
Operation Equipment: Spectrophotometer/Microplate reader
Catalog Number: BC5175
Size: 50T | 48S
Components:
Reagent I: 25 mL×1. Store at 2-8℃.
Reagent II: 6 mL×1. Store at 2-8℃.
Product Description：
Direct bilirubin (DBIL) is also called conjugated bilirubin. After indirect bilirubin enters the liver, it is combined with glucuronic acid by the action of glucuronosyltransferase in the liver. The increase of direct bilirubin is of great significance for clinical diagnosis of obstructive jaundice, hepatocellular jaundice, liver cancer, pancreatic head cancer, cholelithiasis, and cholangiocarcinoma. Direct bilirubin could be oxidized by sodium nitrite to form biliverdin, which has an absorbance of 450 nm. The content of direct bilirubin can be calculated by detecting the wavelength change at 450 nm.
Reagents and Equipment Required but Not Provided：
Spectrophotometer/Microplate reader, desk centrifuge, constant temperature foster box/water bath, pipette, micro glass cuvette/96 well flat-bottom plate, ice, and distilled water.
Procedure
A. Sample preparation:
Serum, plasma, or other liquid samples: Detect the sample directly. If the solution is turbid, measure centrifuging.
B.Determination procedure：
- Preheat the spectrophotometer/microplate reader for 30min, adjust the wavelength to 450 nm, and set the spectrophotometer counter to zero with distilled
- Determination:
| Reagent (μL) | Test tube | Blank tube |
| Sample | 8 | – |
| Distilled water | – | 8 |
| Reagent I | 192 | 192 |
| Mix well. React at 37℃ for 5 minuta. Measure the absorbance at 450 nm, and record it as A1T, A1B; | ||
| Reagent II | 48 | 48 |
| Mix well. React at 37℃ for 5 minuta. Measure the absorbance at 450 nm, and record it as A2T, A2B. Calculate the ΔAT =A1T-A2T and the ΔAB =A1B-A2B. Blank tubes only need to be measured once or twice. | ||
C.Calculations：
96 well flat-bottom plate
DBIL content (μmol/L) =1495.7×(ΔAT-ΔAB)-15.343
Micro glass cuvette
DBIL content (μmol/L) =1099.7×(ΔAT-ΔAB)-10.73
Note:
- Bilirubin decomposes easily in light. Avoid light during
- If the ΔA is lower, it is recommended to increase the sample size before determination; If ΔA＞5, it is recommended to dilute the sample before determination.
Examples:
- Take mouse serum to follow the determination procedure to operate. Determination with 96 well flat-bottom plate, and calculate ΔAT=A1T-A2T=0.217-0.170=0.047, ΔAB=A1B-A2B=0.044-0.044=0. The calculated content is as follows:
DBIL content (μmol/L) =1495.7×(ΔAT-ΔAB)-15.343=54.95 μmol/L.
Povezani proizvodi
BC5180/BC5185 Total bilirubin (TBIL) Content Assay Kit
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