Bacterial Detection of Human Sputum Samples from the Lower Respiratory Tract Using a Microfluidic Chip

To begin, gently shake a thawed isothermal amplification reagent. Centrifuge the reagent to collect all of it at the bottom of the tube. Pipette 20 microliters of the isothermal amplification reagent into a prepared 200 microliter centrifuge tube.

Cover the tube with its lid, and transfer it to the specimen preparation area. Now add 34.5 microliters of the target nucleic acid sample into each tube. Shake gently to mix thoroughly.

Centrifuge the tubes briefly to collect all samples at the bottom of the tubes. Next, write the sample number on a chip packaging label, open the microfluidic chip with the packaging label facing up. Place the chip with the inlet and outlet ports facing upwards.

Then, use a pipette to draw 50 microliters of the prepared amplification reaction system and add it to the main channel of the chip through the inlet port. Stop adding when the main channel is filled. Wipe away any excess liquid around the inlet and outlet ports with lint-free tissue.

Next, click on the open tray button on the nucleic acid analyzer after it is preheated. Place the chip with the front side up on the tray, ensuring that the locating small cylinder emerges from the center gap of the chip to secure it. Press the close tray button to insert the chip into the nucleic acid analyzer.

Enter the sample testing information in the sample information area on the detection interface. Then, press the start detection button to initiate sample detection. Sample one was found to be infected with Klebsiella pneumoniae, while samples two and seven showed polymicrobial infections.

A positive indicator was represented by an amplification curve exhibiting an S shape, while a horizontal line represented a negative indicator.