Instructions for using the apolipoprotein A2 assay kit

The ApoA2 assay kit is mainly used to detect the concentration of ApoA2 in serum or plasma, and its results are of great significance for evaluating lipid metabolism abnormalities, cardiovascular disease risk, etc. The following are the general usage methods and key precautions for the reagent kit. The specific operation should be based on the product manual:

1. Preparation before use

Reagent and sample preparation. The composition of the reagent kit usually includes R1 (buffer), R2 (labeled antibody or enzyme reagent), calibration standard (ApoA2 standard with known concentration), quality control product (used to verify detection accuracy), and diluent (if necessary). Sample requirements: Type: serum (recommended) or plasma (EDTA or heparin anticoagulant).

Collection: Fasting venous blood collection to avoid hemolysis (hemolysis can lead to increased ApoA2 release, resulting in higher results). Storage: Refrigerate at 2-8 ℃ for 3 days, freeze at -20 ℃ for 1 month (avoid repeated freeze-thaw cycles).

Instrument preparation: Calibrate the fully automatic biochemical analyzer or spectrophotometer (wavelength usually 340nm or specific wavelength, depending on the kit).

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Reagent rewarming and mixing: Balance the components of the reagent kit to room temperature (18-25 ℃), gently invert and mix to avoid the formation of bubbles.

. Calibration and quality control samples need to be completely dissolved before use (some test kits require centrifugation to remove sediment).

2. Detection steps (taking immunoturbidimetry as an example)

Method 1: Operation of fully automatic biochemical analyzer

Parameter settings: Set the detection wavelength, reaction temperature (usually 37 ℃), reaction time (such as 5 minutes), and calibration curve type (such as two-point calibration or multi-point calibration) according to the instructions of the reagent kit.

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Enter the concentration value of the calibration standard (such as 0, 50, 100, 200 mg/L, etc.).

Sample addition and detection sample processing: If the sample concentration exceeds the linear range, it needs to be diluted with diluent (such as multiplying the result by 2 after 1:2 dilution).

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Automatic sample addition: The instrument adds R1, sample R2， Incubate the reaction after mixing.

Absorption measurement: After the reaction is complete, the instrument detects the endpoint absorbance or kinetic curve, and calculates the concentration of ApoA2.

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Method 2: Manual operation (spectrophotometer)

Calibration curve drawing: Take 100 μ L of each calibration sample (0, low value, median, high value) and add them to the test tube separately.

. Add 200 μ L of R1 to each tube, mix well, and incubate at 37 ℃ for 5 minutes.

Add 50 μ L of R2, mix immediately, and measure the absorbance (A1) at a wavelength of 340nm after 1 minute. Measure the absorbance again (A2) after 5 minutes.

. Draw a standard curve or calculate a regression equation with the concentration of the calibration standard as the x-axis and Δ A (A2-A1) as the y-axis. Sample detection: Take 100 μ L of sample and measure the Δ A value according to the calibration procedure. Calculate the concentration of ApoA2 in the sample based on the standard curve or regression equation.

III. Quality Control

Quality Control Product Testing: Low and high value quality control products should be tested simultaneously for each batch of testing, and the results should be within the range specified in the instructions (such as low value: 40 ± 5 mg/L, high value: 160 ± 10 mg/L).

. If the quality control results are out of control, it is necessary to investigate the cause (such as expired reagents, instrument malfunctions, sample problems) and retest. Repeatability validation involves testing the same sample three times and calculating the coefficient of variation (CV%), which should be ≤ 5% (usually CV% ≤ 3% using immunoturbidimetry).

IV. Interpretation and Report of Results

Reference Range: Adult Serum ApoA2 Reference Value: 25-50 mg/dL (or 250-500 mg/L, depending on the detection method).

. The reference range for children or special populations (such as nephrotic syndrome patients) may vary and should be combined with clinical judgment.

Increased clinical significance: seen in hyperlipidemia, diabetes, alcoholic liver disease, chronic kidney disease, etc.

Reduction: It is related to the increased risk of cardiovascular disease (ApoA2 is the main component of HDL, with anti atherosclerosis effect). Dynamic changes: ApoA2 levels may improve after treatment interventions (such as statins) and need to be monitored regularly.

V. Precautions

Avoid interfering substances. Hemolysis, hyperlipidemia, and jaundice samples may cause result bias. Centrifuge to remove sediment or dilute before retesting.

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Avoid using anticoagulants containing sodium azide (sodium azide can inhibit enzyme reagent activity).

After opening the bottle, the reagent should be stored at 2-8 ℃ and used within the time specified in the instructions (usually R1 and R2 remain stable for 1 month after opening).

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Avoid exposing reagents to strong light or high temperature environments. Wear gloves and masks during safety protection operations to avoid contact with biological samples and reagents (some reagents contain preservatives such as ProClin, which may irritate the skin). Waste should be disposed of according to biosafety requirements (such as high-pressure sterilization for infectious samples).