A Microfluidic Chip-Based Method for Rapid Neutrophil Chemotaxis Analysis Using Whole Blood
Published: January 31, 2024
Abstract
Source: Yang, K., et al. An All-on-chip Method for Rapid Neutrophil Chemotaxis Analysis Directly from a Drop of Blood. J. Vis. Exp. (124), (2017)
This video demonstrates neutrophil chemotaxis assay using an all-on-chip method. The method allows direct isolation of neutrophils from the whole blood by immunomagnetic negative selection followed by chemotaxis assay on a microfluidic chip.
Protocol
All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines. 1. Microfluidic device preparation. Microfluidic Cell Migration Assay Preparation Prepare 50 µg/mL fibronectin solution by diluting 50 µL of stock fibronectin solution (1 mg/mL) to 950 µL Dulbecco's phosphate-buffered saline (DPBS) inside a biosafety cabinet. Prepare the mig…
Representative Results
Figure 1: Illustration of the all-on-chip method for neutrophil chemotaxis analysis. (A) Illustration of the microfluidic device. The device includes two layers. The first layer (4 µm high) defines the cell docking barrier channel to trap the cells beside the gradient channel. The second layer (60 µm high) defines the gradient generating channel, the port and channel fo…
開示
The authors have nothing to disclose.
Materials
| 装置の製造 | |||
| Mask aligner | ABM | N/A | |
| Spinner | Solitec | 5000 | |
| Hotplate | VWR | 11301-022 | |
| Plasma cleaner | Harrick Plasma | PDC-001 | |
| Vacuum dessicator | Fisher Scientific | 08-594-15A | |
| SU-8 2000 thinner | Microchem | SU-8 2000 | |
| SU-8 2025 photoresist | Microchem | SU-8 2025 | |
| (tridecafluoro-1,1,2,2- tetrahydrooctyl) trichlorosilane | Gelest | 78560-45-9 | |
| Si wafer | Silicon, Inc | LG2065 | |
| Glass slides | Fisher Scientific | 12-544-4 | |
| Cutting pad | N/A | N/A | Custom-made |
| Punchers | |||
| On-chip cell isolation and chemotaxis assay | |||
| RPMI 1640 | Fisher Scientific | SH3025502 | |
| DPBS | Fisher Scientific | SH3002802 | |
| Bovine serum albumin (BSA) | Sigma-Aldrich | SH3057402 | |
| Fibronectin | VWR | CACB356008 | |
| fMLP | Sigma-Aldrich | F3506-10MG | |
| Magnetic disks | Indigo Instruments | 44202-1 | 5 mm in diameter, 1 mm thick |
| FITC-Dextran | Sigma-Aldrich | FD10S | |
| Rhodamine | Sigma-Aldrich | R4127-5G | |
| Giemsa stain solution | Rowley Biochemical Inc | G-472-1-8OZ | |
| EasySep Direct Human Neutrophil Isolation Kit | STEMCELL Technologies Inc | 19666 | |
| Dithiothreitol | Sigma-Aldrich | D0632 | |
| Nikon Ti-U inverted fluorescent microscope | Nikon | Ti-U | |
| Microscope environmental chamber. | InVivo Scientific | N/A | |
| CCD camera | Nikon | DS-Fi1 |
