Label-Free Neutrophil Separation from Tracheal Secretion Using Spiral Microfluidics Channel

Use a 10-milliliter syringe to collect 1 milliliter of airway secretion samples in 9 milliliters of phosphate-buffered saline. Then, with the help of a blunt pipette, homogenize the mucus sample. Next, use a 40-micrometer nylon cell strainer to strain the diluent, and remove large chunks of tissue or blood clots. After the straining is complete, put the sample on ice.

Then, add 50 milliliters of phosphate-buffered saline to the 0.5-milliliter sample diluent to achieve a final concentration of 1,000 times diluted sample. Use the fluid guide to assemble the PDMS chip in order to apply uniform flow to each of the four spiral microchannels. Next, use a male luer connector with 1/16-inch diameter to connect to the inlet, the inner wall outlet, and the outer wall outlet ports of the fluid guide.

Then, connect a silicone tubing to the sample suspension. Next, connect the peristaltic pump to the inlet tubing. Place the outer wall outlet tubing in the waste reservoir. Then position the end of the inlet tubing and the inner wall outlet tubing in the sample reservoir.

After positioning the tubing, leave a 50-milliliter tube filled with 5 milliliters of phosphate-buffered saline without calcium and magnesium in the sample reservoir. Then, in order to prime the device, start pumping at a flow rate of approximately 1 milliliter per minute.

After placing the sample in the sample reservoir, switch on the peristaltic pump and set the flow rate at 4 milliliters per minute. After the sample volume reaches the desired volume of approximately 1 to 2 milliliters, stop the operation and disconnect the tubing.