Mass Photometry to Study Antigen-Antibody Interactions on a Single-Molecule Level
Published: June 29, 2023
Abstract
Source: Wu, D. et. al., Rapid Determination of Antibody-Antigen Affinity by Mass Photometry. J. Vis. Exp. (2021)
In this video, we demonstrate the use of mass photometry to study antigen-antibody interactions in a flow chamber. The focused light scatters differently based on the mass of the molecules. A detector collects the scattered light and generates the interference pattern as spots of varied intensities, which correlate with the molecular masses.
Protocol
1. Prepare the flow chambers Clean the glass coverslips Using wash bottles with distilled water, ethanol, and isopropanol, rinse the 24 mm x 50 mm coverslips in the following order: water, ethanol, water, isopropanol, water. Dry the coverslips with a stream of clean nitrogen. It is important to rinse the coverslips from top to bottom, holding the bottom corner with soft-tipped forceps. Dry the coverslip in the same direction to avoid transferring contamination from the forceps (<str…
Representative Results
Figure 1: MP flow chamber preparation and loading. (A) Coverslip holding position for the cleaning procedure. (B) Alignment of the 24 x 24 mm coverslip (middle layer) and the double-sided tape (top layer) on the surface of aluminum foil (bottom layer, not shown). Blue dashed lines show the location of the cut lines. (C) Top and side view of …
開示
The authors have nothing to disclose.
Materials
| AcquireMP | Refeyn | MP data collection software | |
| Anti-human thrombin | Haematologic Technologies | AHT-5020 | RRID: AB_2864302 |
| Cotton-tipped applicators | Thorlabs | CTA10 | Cotton optical swabs for lens cleaning |
| Coverslips 24×24 mm | Globe Scientific | 1405-10 | |
| Coverslips 24×50 mm | Fisher Scientific | 12-544-EP | |
| DiscoverMP | Refeyn | MP data processing software | |
| Forceps | Electron Microscopy Sciences | 78080-CF | Soft-tipped forceps for coverslips handling |
| Human α-thrombin | Haematologic Technologies | HCT-0020 | |
| Immersion oil | Thorlabs | MOIL-30 | |
| Isopropanol | Alfa Aesar | 36644 | |
| Microsoft Excel | Microsoft | Spreadsheet | |
| OneMP | Refeyn | Mass Photometry instrument | |
| Origin | OriginLab | Scientific graphing software | |
| PBS | Corning | 46-013-CM | 10x stock |
| Syringe filter | Millipore | SLGSR33SS | Buffer and sample filtering |
