Three-Dimensional Quantitative Phase Imaging for Characterizing Lymphocyte Subtypes
Published: November 30, 2023
Abstract
Source: Yoon, J. et al. Label-Free Identification of Lymphocyte Subtypes Using Three-Dimensional Quantitative Phase Imaging and Machine Learning. J. Vis. Exp. (2018)
This video demonstrates label-free identification of lymphocyte morphology and biochemistry through 3D quantitative phase imaging. This technique allows for a thorough analysis of lymphocytes, uncovering their internal structures and properties without the need for labeling procedures.
Protocol
1. Flow Cytometry and Sorting of Lymphocyte Subtypes NOTE: Sorting lymphocytes depending on cell type is essential for establishing the ground-truth (i.e., correct) cell type labels to train and test a cell type classifier in supervised learning. Flow cytometry, a gold standard method, is used to identify and separate lymphocytes. Make a mixture of surface marker staining antibodies in 100 µL of fresh RPMI-1640 medium [with 10% FBS, 0.1 µg of CD3…
開示
The authors have nothing to disclose.
Materials
| Flow cytometry | BD Biosciences | Aria II or III | |
| Imaging chamber | Tomocube, Inc. | TomoDish | |
| Holotomography | Tomocube, Inc | HT-1H | |
| Holotomography imaging software | Tomocube, Inc. | TomoStudio | |
| Image-processing software | MathWorks | Matlab R2017b | |
| Falcon conical centrifuge tube | ThermoFisher Scientific | 14-959-53A | 15 mL |
| Phosphate-buffered saline | Sigma-Aldrich | 806544-500ML | |
| Ammonium-chloride-potassium lysing buffer | ThermoFisher Scientific | A1049201 | |
| RPMI-1640 medium | Sigma-Aldrich | R8758 | |
| Fetal bovine serum | ThermoFisher Scientific | 10438018 |
