First, the choice of microplate reader Faced with such a large number of microplate readers, the laboratory often has difficulty in adapting when choosing. In general, the microplate readers used for enzyme immunoassay can be divided into common microplate readers, multifunctional microplate readers and fully automated enzyme immunoassay systems. The function of common microplate readers is basically equivalent to L-plate colorimeter, and multi-functional automatic microplate reader in addition to the microplate colorimetric function, there are enzyme labeling kinetics, UV and even agglutination and other detection functions; automatic enzyme immunoassay system has The "open" and "restricted" reagents, the so-called reagent "open" means that different brands of ELISA reagents can be used for the instrument, and the reagent "limited" must use the instrument-specific enzyme-free assay reagents. When selecting a microplate reader, laboratories should generally follow these principles:
1. According to work needs to choose. Do not deliberately seek more functions, because one instrument if the function is too much, easy to fail, and some other functions such as enzyme kinetics or agglutination, may not be used at all; if the intended use of microplate reader is mainly used For qualitative determinations, there is no need for a perfect software system for quantification. As for the automatic enzyme immunoassay system, it is more suitable for blood stations or large hospitals with a large workload, and it is not necessary for laboratories with less daily workload.
2. According to the performance of the microplate reader to choose. The performance indicators of the reaction microplate reader mainly include the measurement wavelength range, filter configuration, detection speed, absorbance range, linear range, resolution, accuracy, and precision.
3. According to the financial resources of your laboratory to choose. There are many kinds of microplate readers, because the performance is not the same, the price will naturally differ greatly, the laboratory can choose the appropriate microplate reader based on their own financial resources.
4. Choose according to after-sales service. After the laboratory instruments are purchased, just like the household appliances used in our homes, during use and after a certain period of use, there is likely to be warranty or maintenance issues, so be sure to choose the enzymes of the company with good after-sales services. Marker.
5. Choose according to the foreign language level of laboratory personnel. Many of the existing microplate readers have used Chinese man-machine conversations. For labs with difficulties in English, such microplate readers should be used as much as possible.
After determining the selection principle, it is how to choose. The first step in the selection process is to obtain information about the microplate reader. There are generally several ways to do this:
1 Consultation with a number of laboratory colleagues who have purchased different microplate readers to obtain the most direct information on their use.
2 Microplate reader seller's information advertisement and introduction;
3 The instrument evaluation report of the authoritative organization.
After obtaining as many microplate reader information data as possible, it is to compare the performance indicators of different types of microplate readers, and to select one or two or three more suitable for the laboratory, and then make further detailed consultation to the seller. Learn to decide which one to choose for self inspection.
Second, the microplate reader self-checking and calibration from the above information obtained by the microplate reader performance information, basically can be said to be indirect, with the goal, if you still want to obtain a perceptron understanding of a microplate reader , It can be self-tested, mainly in the following aspects.
(1) Appearance The microplate reader should have the instrument name, model number, serial number, manufacturer, date of manufacture, and power supply voltage; the adjustment knobs, buttons, and switches can all work normally; the outer surface is free from mechanical damage; the display text should be clear and complete. .
(b) Stability of the microplate reader (drift)
Choose 492nm wavelength, in absorbance o. At 0A, measure the spectral neutral filter with nominal value of 1.0A (measurement operation according to the instrument instructions), record the instrument indication or average value, record the instrument indication or average value again after 10 minutes, and measure the value twice before and after The difference should not exceed ±0.005A.
(3) Accuracy of absorbance measurement Select wavelengths of 405, 492 and 620 nm. Using air as the reference, determine the spectral neutral glass filters with nominal values ​​of 0.5 and 1.0 A, respectively, and replace them with special test plates. The ELISA plate was continuously measured 3 times according to the instrument instructions. The accuracy was calculated according to the following formula: AA = 1/3 (A, +A. + A3) - A: (A: is the standard value).
(4) The repeatability wavelength of absorbance determination is the same as (3). A blank solution is filled in the first row of the ELISA plate, and a potassium dichromate solution with a concentration of 200 rig/ml is added in the second row and the determination is repeated 5 times. Record the absorbance of the second row of each measurement and take the absorbance of any well in the second row and absorbance of each well. Calculate the repeatability as follows:
(e) The linearity of the microplate reader is 540nm and the nominal value is o. 5, 1.0A neutral filters were placed in the sample position of the special test board, with air as the reference, and each measurement was repeated 3 times; then the above two neutral filters were superimposed and placed on the dedicated test board. In the same well position, the measurement was repeated 3 times. Linear error:
Where A1 is the average absorbance of the first neutral filter, A: is the average absorbance of the second neutral filter, and A1 and 2 are the absorbance averages of the two neutral filters.
(six) test speed determination.
A wavelength of 492 nm was used, and a 96-well microtiter plate was placed for measurement. The stopwatch was used to measure the time at which the instrument printed out all the data.
The microplate reader should be self-checked as far as possible in addition to the purchase, and it should also be periodically tested during use to ensure the validity of the microplate reader assay.
Attachment: Preparation of 200ttg/ml potassium dichromate solution Place potassium dichromate solid reagent in a weighing bottle (decap) in an oven, bake at 105±5°C for 2h, and place in a desiccator to cool 30min, accurately weighed on the analytical balance o. 2829g, placed in a 100ml beaker, with o. Diluted sulfuric acid 05mol / L, transferred into a 500ml volumetric flask, with a small amount of o. 05mol/L dilute sulfuric acid washing beaker 3 times, washing liquid into the volumetric flask, diluted with 0.05mol/L dilute sulfuric acid to the mark, shake.