C peptide radioimmunoassay kit instructions

C-peptide radioimmunoassay kit manual [drug name]

Common name: Iodine [125I] C peptide radioimmunoassay kit

English name: Iodine [125I] C-peptide Radioimmunoassay Kit

Chinese Pinyin: Dian [125I] C-tai Fangshemianyifenxi Yaohe

C peptide radioimmunoassay kit manual [ingredients]

【Properties】 Liquid components (except immunoseparating agents) should be clear, without precipitation or flocs; freeze-dried components should be loose, and after reconstitution, there should be no sediment or flocs.

[Half-life of radionuclide] The nuclide used in this product is iodine [125I], and its physical half-life (T½) is 60 days.

【Clinical Significance】

C peptide (C-Peptide, CP) is a linear polypeptide consisting of 31 amino acids with a molecular weight of 3021. Under the action of invertase, proinsulin produces equal moles of C-peptide and insulin. Since the C peptide and insulin are released from the insulin cells to the blood in an equimolar ratio, the measurement of serum C-peptide can reflect the function of islet β cells; in addition, because insulin antibodies and C-peptide do not cross reflect, and medicinal insulin does not contain C peptide, so exogenous insulin does not affect the radioimmunoassay of C peptide. Therefore, the determination of serum C-peptide can more accurately judge the secretory function of β cells for species receiving insulin therapy. Moreover, it is of great significance for the classification of diabetes, treatment and prognosis estimation, as well as research on diseases related to glucose metabolism.

This kit is mainly used to determine serum CP content.

【Measurement principle】

This method uses competitive radioimmunoassay, that is, CP and 125I-CP in the standard or test sample together with a limited amount of specific CP antibody under competitive conditions for competitive binding reaction. A part of 125I-CP binds to the antibody to form a complex, while the other part is in a free state. The ratio of 125I-CP to antibody depends on the content of unlabeled CP in the standard or the sample to be tested. The higher the content of unlabeled CP, the less complex 125I-CP forms with the antibody, the lower the content of unlabeled CP -The more complexes the CP forms with the antibody. The antigen-antibody complex is precipitated with an immunological separating agent, and the radioactive count of the precipitate is measured. After proper data processing, a standard curve can be obtained, and the CP content of the tested sample can be found from the standard (the data can be automatically processed by the computer).

C-peptide radioimmunoassay kit manual [applicable instrument] is applicable to all kinds of γ-radioimmunoassay analyzers.

[Sample requirements]

Take venous blood to separate serum for testing; if not to measure immediately, the sample can be sealed and stored at -20 ℃ for testing, and stored at -20 ℃ for no more than one month. Frozen samples should be protected from repeated freezing and thawing during storage. Before measuring the frozen sample, let the sample melt in room temperature or normal temperature water and shake it well.

【Dosage】

1 Reagent preparation, usage and storage

(1) CP standard product: lyophilized product, add 1ml of buffer to each bottle of standard product to dissolve. Shake well after dissolving to apply. After dissolution, the standard concentration values ​​were 0.25, 0.5, 1.0, 3.0, 6.0, 12.0ng / ml. After standard reconstitution, it can be stable for 42 days when stored at -20 ℃, and the freeze-dried product is stable for -20 ℃.

(2) 125I-CP: lyophilized product, dissolved in buffer, add 5ml per bottle, shake well before adding sample. The radioactivity of each bottle of marker is not more than 74KBq (2μCi), and the unused marker can be stored for 42 days at -20 ℃.

(3) Rabbit anti-CP antibody: freeze-dried product, dissolved in buffer solution, add _ml per bottle of antibody (if not filled in 100T plus 10ml, 50T plus 5ml buffer solution). After the antibody is dissolved, it can be stable for 42 days when stored at 2-8 ° C, and the lyophilized product is stable during the storage period of -20 ° C.

(4) Donkey anti-rabbit immunoseparation reagent: liquid, it must be shaken well before loading! Store at 2 ～ 8 ℃, stable within the validity period. Stable within 45 days after opening.

(5) CP buffer solution: liquid, stored at 2 ～ 8 ℃, stable within the validity period.

(6) CP quality control serum: freeze-dried product, add 0.5ml of distilled water to each bottle of quality control serum before use to dissolve. After reconstitution, storage at -20 ° C can be stable for 42 days, and lyophilized products are stable at -20 ° C for the period of validity. For the validity period and value-added, please refer to the applicable instructions of the quality control serum.

C peptide radioimmunoassay kit instruction manual 2 Measurement steps

Take a number of round bottom polystyrene test tubes, use a marker or special pencil to number NSB, S0 ~ S6 and the sample tube to be tested, etc., then use a micro sampler to add samples according to the table below. Before loading, all reagents (especially the separating agent!) Including the sample to be tested should be shaken, and it is best to equilibrate to room temperature.

Sample loading program unit: μl

3 Data processing

(1) Online processing: the result is automatically processed by the computer. Note that due to different principles of radioimmunoassay and immunoradiotherapy, the processing software is also different. Users must use suitable processing software for data processing. It is recommended that users use logit-log or four-parameter data processing mode.

(2) Hand drawing or calculator processing:

1). Percentage binding rate calculation: S0 tube count is B0, each standard tube or sample tube count is B, non-specific tube count is NSB, then the percentage binding rate calculation formula is: B / B0 = (B－NSB ) / (B0－NSB) × 100%

2). Logit calculation: The formula for calculating the logit value of each standard point or sample tube is: logit = ln (B / B0) / (1－B / B0)

3). Hand work diagram: take the log value of the standard concentration as the abscissa, the corresponding logit value as the ordinate on the ordinary coordinate paper or the standard concentration as the abscissa, and the corresponding B / B0 as the ordinate in the logit-log coordinate Draw a standard curve (a straight line when idealized) on paper. According to the B / B0 of the sample to be tested, the concentration value of the sample can be found from the coordinate paper. Note: If you use ordinary graph paper, the detected value should take the logarithm to be the final concentration value.

4). Calculator processing: using a calculator with linear fitting function, the sample concentration value can be calculated more conveniently, please refer to the instruction manual of the calculator for the specific operation method.

Examples of standard curves (for reference only, not for actual calculations).

【Refer to normal value】

The fasting serum CP reference normal value: average 1.6ng / ml, range 0.6 ～ 3.4 ng / ml.

The peak value of serum CP after a meal is about 5 times that of a fasting. The peak generally appears 30 to 60 minutes after a meal (it has a certain relationship with the type of food eaten), and it is close to the fasting value after 3 hours.

Unit conversion: 1ng / ml = 100ng / dl = 1000ng / l = 1μg / l = 0.331nmol / l

Due to different regions and instruments used, there will be some differences in the measured normal values, so the normal values ​​provided in this manual are for reference only. It is best for each laboratory to establish a range of normal values ​​that meets the requirements of this laboratory.

【Product Performance Index】

1. Measuring range: 0.25 ～ 12ng / ml

2. Sensitivity 0.1 ng / ml

3. Precision: intra-batch coefficient of variation (CV) <10%; inter-batch coefficient of variation (CV) <15%.

4. Term of use: See product packaging.

[Taboo] This product is only for in vitro diagnosis!

【Precautions】

1. After receiving the kit, store it at 2 ～ 8 ℃ as soon as possible.

2. Please read this manual carefully before use.

3. Standards and markers are easy to be inactivated after dissolving. After dissolving, they should be stored at -20 ℃ in time and divided into several -20 ℃ if necessary.

4. The components of the same kind of kits in different kits or in different months must not be mixed.

5. When aspirating the supernatant, be careful not to lose the sediment, otherwise it will significantly affect the measurement results.

6. When the CP concentration in the sample is higher than the concentration at point F, the sample can be diluted 5-10 times with buffer and then retested. The measured concentration multiplied by the dilution factor is the sample concentration.

7. Please use the kit within the validity period.

8. Clinical specimens should be regarded as potentially infectious. Please follow the laboratory procedures for infectious diseases.

9. The use of this kit should pay attention to radiological protection, and radioactive waste must be disposed of in accordance with the regulations on radiological protection.

10. Operators using this product should undergo professional technical training.