Animal disease rapid detection all-in-one machine
Animal Disease Rapid Detection All-In-One Machine
Operating Instructions
The use of the animal disease rapid detection all-in-one machine mainly includes four modules：
一、 Preliminary Treatment Of Diseased Materials
二、 Nucleic Acid Extraction
三、 Gene Amplification
四、 Result Analysis
Step 1: Preliminary Treatment of Diseased Material
Types of Animal Disease Materials: Animal disease materials primarily consist of serum, cotton swabs (oral / nasal swabs, cloacal swabs, etc.), and tissue samples (heart, liver, spleen, lung, kidney, trachea, etc.), etc. Serum and cotton swabs can be centrifuged in a handheld centrifuge for 1 minute to proceed to nucleic acid extraction (Module 2) without grinding.
Steps for Handling Animal Tissue Samples:
1、Remove the grinder head and open the baffle.
2、 Grinding: Use scissors to place the tissue sample into the grinder. Add sterile saline or purified water (approximately 3 times the tissue volume). Cover the baffle, attach the grinder head, and press to start grinding (1-2 minutes).
Note: Intermittent pressing ensures thorough grinding.
3、 Transfer: Remove the grinder head and open the baffle. Use a 200 μL pipette with a yellow tip to aspirate the ground sample 3-5 times into a 1.5 mL centrifuge tube.
Note: Yellow tips are disposable; avoid reuse.
4、 Centrifugation: Turn on the handheld centrifuge, place the 1.5 ml centrifuge tube containing the diseased material in a handheld centrifuge with a balanced tube (same volume of water). Cover the machine, and centrifuge at 3-5 minutes. The supernatant is the processed animal tissue diseased material, which is used for nucleic acid extraction in the second module.
Note: Handle the 1.5 ml centrifuge tubes gently to avoid remixing layers.
Step 2: Nucleic Acid Extraction
1、 Prepare the Extraction Reagent Sleeve: Open the viral DNA/RNA purification kit, take out the extraction reagent sleeve, tear off the sealing film, and insert the sleeve into the deep-well plate (tear the seal first).
Notes：The leftmost well contains black magnetic beads. Avoid inserting the deep-well plate too deeply to ensure proper contact with the heating block.
2、 Add Sample:: Install the yellow pipette tip on the 200μl quantitative pipette, and pipette 200 μL of supernatant into the first left well (black beads).
Note: Change tips between samples to prevent cross-contamination.
3、 Power On: Turn on the power switch on the back of the all-in-one machine, and the machine will automatically turn on. When entering the "动物疫病快速检测一体机" interface, click the "进入系统" button to enter the all-in-one machine operation interface.
4、 Load Magnetic Sleeves: Click the "提取" button in the upper left corner of the operation interface to enter the extraction interface, then click the "装磁套" button in the upper right corner, the motor at the bottom of the all-in-one machine starts, and the magnetic rod automatically moves to the magnetic rod sleeve unloading position. Open the door of the nucleic acid extraction cabin, insert the magnetic rod sleeve to the bottom, click the "装磁套" button in the upper right corner, and the magnetic rod automatically resets.
5、 Start Extraction: Place the mold deep well plate to be tested in the instrument slot, with the hole with black magnetic beads on the left side of the slot and press the extraction reagent sleeve to prevent poor contact between the sleeve and the heated aluminum block.
Close the door, click the "开始" button on the right side of the extraction interface, enter the start confirmation interface, click the "是" button, and the nucleic acid extraction begins.
6、 Completion: The entire nucleic acid extraction process takes about 25 minutes. When the pop-up window of the experiment completion appears, click the "确认" button and the nucleic acid extraction ends. Open the door and take out the mold deep well plate. Click the "装磁套" button on the extraction interface, the motor at the bottom of the integrated machine starts, the magnetic rod automatically moves to the magnetic rod sleeve unloading position, pull out the magnetic rod sleeve, click the "装磁套" button again, the motor resets, closes the hatch, and the extraction module is returned to its original position.
The eluate in Well 6 (with the black magnetic bead hole as the first hole) is the extracted nucleic acid, which can be directly used for gene amplification in the third module.
Step 3, Gene Amplification
1、Preparation: Select the appropriate fluorescent PCR reagent for the target disease.
Note:
The virus fluorescent PCR amplification reagent can be cut and used according to the number of samples to be tested. Do not open the unused reagent to prevent contamination.
To prevent the amplification reagent from sticking to the tube wall and insufficient fusion, you can flick the reagent tube a few times with your hand before adding the nucleic acid.
2、Install the yellow pipette tip on a 25μl quantitative pipette, respectively absorb the nucleic acid to be detected, and add it to the corresponding virus fluorescent PCR amplification reagent reaction tube. Replace the yellow pipette tip when aspirating different nucleic acids to prevent cross contamination. Tighten the PCR reaction tube cover and mark it.
Open the handheld centrifuge, remove the round rotor of the handheld centrifuge, replace it with the square rotor of the PCR tube, put the PCR reaction tube to be tested, and balance it with PCR tubes of the same weight in symmetrical positions (please use the PCR balancing tubes that come with the machine when there are odd numbers, and self-balance them in symmetrical positions when there are even numbers).
Close the machine cover, and the centrifugal time is about 1-2 minutes (note: not less than 1 minute!). Press the top cover switch of the all-in-one machine to open the PCR amplification chamber. Put the PCR reaction tube to be tested into the PCR amplification hole and close the door.
Note：
The all-in-one machine has a total of 8 PCR amplification wells, from left to right, wells 01 to 08, and each well can hold one PCR reaction tube.
The well number of each diseased material and its corresponding reagent part number must be recorded.
3、Program Setup: Click the "Amplification" button on the operation interface to enter the amplification module interface. Click the "Start Experiment" button to enter the "Create Experiment" interface.
Note：“
Note: The "扩增程序" numbers (left side of the "创建实验") match the reagent product codes on the PCR kit box.
The number on the left side of the "创建实验" interface represents the "amplification program". Select the "amplification program" with the same number according to the reagent placed in the No. 01 hole of the integrated machine, click the area below the well information "1" to select the "amplification program", and then click the "确定" button on the right to complete the association setting between the No. 01 hole of the integrated machine and its "amplification program". Similarly, complete the association setting between other wells and their "amplification programs".
4、Start Amplification: Click the "开始" button to start gene amplification, enter the start confirmation interface, and click "开始" to start gene amplification.
5、End Amplification: The entire gene amplification process takes about 55 minutes. When the pop-up window of the experiment completion appears, click the "是" button, the gene amplification is completed, and you can directly enter the result analysis of the fourth module.
Step 4: Result Analysis
View Test Results: In the result analysis interface, select parameters such as well position and channel to view the test results of the corresponding diseased material.
The all-in-one machine is designed with four fluorescent channels, that is, each PCR amplification well can detect four different diseases at the same time, and one gene amplification can detect up to 32 diseases at the same time.
Test results analysis
Please be informed before the analysis:
The machine supports 4-channel detection per well (8 wells total), enabling simultaneous testing for up to 32 diseases.
Four fluorescent channels (FAM, HEX, ROX, Cy5) correspond to the corresponding diseases of different diagnostic kits. Please refer to the instructions of the kit for details.
CT value indicates the cycle threshold for fluorescence detection. Lower values = higher viral load.
1、 When the PCR amplification test is completed, a pop-up window will appear asking “PCR运行完成，是否进行结果分析？” Click “是” to enter the result analysis interface.
2、The result analysis interface is divided into four areas:
The lower left is the "well position information" area: the space below the number in the well position information corresponds to the PCR amplification well position of the integrated machine, such as the space below the number "1" corresponds to the first well position 01 on the left side of the PCR amplification well of the integrated machine, the space below the number "2" corresponds to the second well position 02 on the left side of the PCR amplification well of the integrated machine, and so on. You can click the space below the number in the "well position information" to select the well position where our sample to be tested is placed and view the test results of the corresponding sample to be tested.
The middle part below is the "Channel Selection" area: Our all-in-one machine is designed with four channels, and each channel corresponds to one of the diseases in different diagnostic kits. You can view the disease test results represented by it by selecting one of the channels "FAM, HEX, ROX, Cy5", or you can view the results of the four diseases by selecting all of them. Generally, unless there are special circumstances, all four channels can be selected for easy viewing of the results.
The upper right corner is the detailed result display area: This area details the hole information of the sample to be tested, the test items, the test diseases, the test results and the specific CT value, etc. The detailed description is as follows:
The first column "孔号" consists of two digits, which are consistent with the numbers in the "Hole Position Information" area on the lower left.
The second column "项目" is consistent with the reagent product number of the PCR amplification tube, indicating which PCR amplification reagent is used for the sample to be tested in this "well number".
The third column "基因" indicates which disease is detected by the PCR amplification reagent used in the sample to be tested in this "well number". Diseases are generally represented by English abbreviations, and the specifics can be confirmed by checking the instructions of the PCR amplification reagent used.
The fourth column "结果" indicates the positive or negative result of the sample to be tested in the "well number" for the disease, which is displayed in red "positive" and black "negative" respectively.
The fifth column "CT值" indicates the specific CT value of the disease corresponding to the sample to be tested in the "well number". The smaller the CT value, the higher the virus content. CT value>38 is negative, and CT value≤38 is positive.
The upper left is the amplification curve area: here you can judge whether the disease detection is normal by observing the curve. The normal amplification curve is an "S"-shaped smooth curve. The colors of the four channels are different and the results do not interfere with each other. The FAM channel is a blue curve, the HEX channel is a green curve, the ROX channel is a light pink curve, and the CY5 channel is a red curve.
3、 Example of result analysis
1) Diseased material: liver of diseased duck.
2) Disease to be detected: diseases related to duck viral hepatitis.
3) PCR amplification reagent: product number HY-KZ-03, where FAM represents DHAV1 (duck viral hepatitis type 1); HEX represents DHAV3 (duck viral hepatitis type 3); ROX represents DAstV1 (duck astrovirus type 1); CY5 represents DPMV (duck paramyxovirus).
4) PCR detection well: well 02.
First, click the space below the number 2 in "孔位信息", select all four channels in "通道选择", and then slide the slider into the upper right result display area to view the results. The results show that the diseased material is positive for duck astrovirus type 1.
5) View historical records
Click any of the other four options (original curve, calibration curve, temperature setting, real-time temperature) on the "扩增" interface except "结果分析", then click "文件管理" in the lower right corner, select the historical record file you want to view in the pop-up dialog box, and click "确定" to view it.