Luminescent immunoassay technology is a method of combining luminescent reaction with immune reaction to detect antigens or antibodies. It uses micro-multiplication technology with good sensitivity and specificity; the detection range is very wide, ranging from traditional proteins, hormones, enzymes to drugs. The most commonly used clinical instruments are fully automatic chemiluminescent immunoassay analyzers, fully automatic microparticle chemiluminescent immunoassay analyzers and fully automatic electrochemical luminescent immunoassay analyzers.
Basic structure of luminescent immunoassay analyzer
1. Composition of fully automatic chemiluminescent immunoassay analyzer
1) Main unit is the operating reaction measurement part of the instrument, including raw material equipment, liquid circuit, mechanical transmission, optical path detection, and circuit part.
The raw material equipment part includes reaction cups, sample trays, reagent trays, pure water, cleaning fluid, and wastewater storage and treatment devices on the machine; the liquid circuit part includes filters, sealing rings, vacuum pumps, pipelines, sample and reagent probes, etc.; the mechanical transmission part includes sensors, transport tracks, etc.; the optical path detection part includes light sources, spectroscopic devices, photomultiplier tubes; the circuit part includes power supply and amplification processing system and circuit control board.
2) The microcomputer processing system is the key part of the instrument and is the command and control center.
Its functions include program-controlled operation, automatic monitoring, indication judgment, data processing, fault diagnosis, etc., and it is equipped with a CD. The host is also equipped with a reserved interface, which can automatically process other data through an external storage device and remotely operate it, which is used for the extension and development of laboratory automation.
2. Composition of the fully automatic microparticle chemiluminescence immunoassay analyzer
1) Sample processing system includes the transfer cabin and the main probe system, which is responsible for adding specimens, reagents, and buffers to the reaction tube.
2) Experimental operation system is the fluid system, which consists of flushing liquid, waste liquid, substrate pump and valve, vacuum pump, water storage tank, liquid tank and probe flushing tower.
3) Central supply and control system consists of reaction tube bracket, reaction tube supply cabin, constant temperature belt and photoelectric reading cabin. It is responsible for transmitting the reaction tube, and during the transmission process, the reaction tube is heated to a certain temperature through the constant temperature belt. When the constant temperature process is completed, the photoelectric recognition device converts the optical signal into an electrical signal.
4) Microcomputer control system consists of a printed circuit board, power supply, hard disk drive, floppy disk drive, restart button and internal lock switch. Peripheral equipment includes color monitor, printer, keyboard, external barcode recognition pen, external barcode scanner and connecting arm. The instrument can be operated by corresponding instructions and data can be read and archived.
3. Composition of fully automatic electrochemiluminescence immunoassay analyzer
It is mainly composed of sample tray, reagent kit, incubation reaction tray, electrochemical detection system and computer control system, which can be divided into three unit modules.
1) The control unit is a complete computer, equipped with a bracket and printing system.
2) The core unit is mainly composed of barcode reader, specimen compartment, specimen rack turntable, module track, etc.
3) The analysis module is the core of the detection system, mainly including pre-cleaning area, measurement area, system reagent area, reagent area, and consumables area.
Working principle of luminescent immunoassay analyzer
Chemiluminescence immunoassay technology is also called micro-multiplication technology, including two methods:
Competitive method: mostly used to determine small molecule antigen substances. An excess of antibodies coated with magnetic particles are added to the reaction cup for incubation together with the antigen to be tested and a quantitative labeled acridinium ester antigen, so that the labeled antigen and the antibody (or the antigen to be tested and the antibody) combine to form a complex.
Sandwich method: mostly used to measure macromolecular antigen substances. The labeled antibody and the antigen to be tested combine with the coated antibody at the same time to generate a complex of the coated antibody-luminescent antibody of the antigen to be tested. The instrument uses certain chemical groups to mark the antigen or antibody. After the chemical group is oxidized, it forms an excited state and releases photons of a certain wavelength in the process of returning to the ground state. The photomultiplier tube converts the received light energy into electrical energy, reflects the light measurement in digital form, and then calculates the concentration of the measured object.
Fully automatic microparticle chemiluminescence immunoassay
Applying the classic immunological principle, monoclonal antibody reagents are used, magnetic particles are used as solid phase carriers, alkaline phosphatase is used as the standard, and the luminescent agent is 3-(2'-spiroadamantane)-4-methoxy-4-(3"-phosphoryloxy)benzene-1,2-dioxetane (AMPPD). Small molecules are determined by competition or antibody capture, while macromolecules are determined by sandwich method.
Fully automatic electrochemiluminescence immunoassay
The sample to be tested is mixed with paramagnetic particles coated with antibodies and antibodies labeled with luminescent agents in a reaction cup and incubated together to form a microbead-coated antibody-antigen-luminescent agent-labeled antibody complex. When the magnetic particles flow through the electrode surface, they are attracted by the magnet installed under the electrode, and the free luminescent agent-labeled antibody is washed away by the buffer. At the same time, a voltage is applied to the electrode to make the luminescent marker terpyridine ruthenium [Ru( bpy)3]2+ transfers electrons on the electrode surface, generating electrochemical luminescence, and the intensity of the light is proportional to the concentration of the antigen to be tested. The inhibition immunoassay is used for the detection of small molecular weight protein antigens; the sandwich immunoassay is used for the detection of large molecular weight substances.
Features of the luminescent immunoassay analyzer
It combines chemiluminescence technology with magnetic microparticle separation technology. It is a fully automatic, random access, software-controlled intelligent analysis system. In the reaction system, magnetic particles are used as solid phase carriers. Its diameter is only 1.0um, which greatly increases the coating surface area, increases the adsorption of antigens or antibodies, speeds up the reaction, and makes cleaning and separation easier. It has the advantages of flexible operation, accurate and reliable results, long reagent storage time, and high degree of automation.
Fully automatic microparticles Micro-particle chemiluminescence immunoassay
Uses microparticle chemiluminescence technology to quantitatively determine trace components and drug concentrations in the human body, with high specificity, sensitivity and stability.
Fully automatic electrochemiluminescence immunoassay
Electrochemiluminescence immunoassay is a specific chemiluminescence reaction triggered by electrochemistry on the electrode surface, which belongs to the third generation of chemiluminescence immunoassay technology. Compared with other immunoassay technologies, it has very obvious advantages: because the marker used, terpyridine ruthenium, can be combined with various compounds such as proteins, hapten hormones, nucleic acids, etc., the detection items are very wide; because the magnetic microbead coating adopts the new solid phase coating technology of "streptavidin biotin", the detection sensitivity is higher, the linear range is wider, and the reaction time is shorter .
Maintenance of luminescent immunoassay analyzer
Advanced equipment requires proper maintenance. Daily maintenance, weekly maintenance and regular system testing are the prerequisites for ensuring the normal operation of the instrument. The maintenance of the fully automatic chemiluminescent immunoassay analyzer includes the following aspects.
1. Daily maintenance: Keep the machine casing clean every day to prevent dust from entering the instrument. Before doing routine maintenance, be sure to check whether the system temperature status, liquid circuit part, consumables part, waste liquid tank, buffer solution, etc. all meet the requirements, and then clean the system according to the maintenance procedure for maintenance operations.
2. Weekly maintenance: Check the upper rail of the main probe, clean the lower rail of the main probe with a fiber-free swab after the inspection, and then enter the maintenance program under the main menu for special cleaning as required. After cleaning, clean the upper part of the main probe with an ethanol swab, and then check the waste night tank filter. Check whether there is dust at the sensing point on the incubation belt and wipe it clean with a fiber-free swab. Be sure to do a system test after weekly maintenance to ensure that the system test data is within the control range.
3. Monthly maintenance: Use a special stainless steel small brush to brush the inside of the main probe, specimen sampling needle, and reagent needle once a month to remove dirt. Since the internal space of the needle is small, after brushing, use a syringe to draw physiological saline to repeatedly rinse the inside of the needle to flush out all the dirt. The outside of the needle can be wiped clean with alcohol [1].
Common troubleshooting of luminescent immunoassay analyzers
Chemiluminescent immunoassay analyzers have a high degree of automation and are equipped with self-diagnosis functions. Once a fault occurs, the instrument can generally detect it automatically, display an error message and sound an alarm.
Common faults mainly include the following aspects.
1. The pressure gauge indicates zero: During a vacuum pressure test, the sound of the pump can be heard, but the pressure gauge indicates zero. First, check the vacuum tube connected to the waste liquid bottle and test the vacuum pressure to determine whether the fault is caused by leakage or damage to the pressure gauge. Check the interfaces of each pipeline for leakage and the four related solenoid valves (during the vacuum pressure test, the four solenoid valves do not work and are in a closed state). Repair or replace the problematic pipelines or solenoid valves in a timely manner.
2. Insufficient vacuum pressure: Perform a vacuum pressure test. If the test result is normal, it can be known that the vacuum sensor cannot detect the vacuum pressure. The pressure test of this machine is to detect high and low pressures by two sensors respectively. After adjusting or cleaning the problematic sensors, the vacuum pressure is tested again. After the pressure is normal, the sensor screws are adjusted to make the high and low pressure indications within the specified range.
3. Luminous body error: Check the surface of the luminous body and find that there is liquid seepage. This fault is checked in three steps: check whether the waste liquid probe, related pipelines and cleaning pool are blocked or leaking; check the sample addition solenoid valve, drain solenoid valve, solenoid valve, and dirt will cause water inlet or drainage; check whether the alkali pump cleaning pipeline connected to the waste liquid probe pipeline is leaking and whether the alkali pump has cracks.
4. Track error: This fault is caused by the misalignment of the specimen rack in the track, making the track unable to run. Because the track is very long and closed and not easy to disassemble, generally check the horizontal elevator connected to the track first. If it is normal, check the track again. As long as the misaligned specimen rack is taken out, the fault can be eliminated.
