Flow cytometry, cell sorting and fluorescence microscopy for infectious disease research
The LIDR Immunology core offers flow cytometry, cell sorting and fluorescence microscopy services for fixed or live BSL-1, BSL-2, and BSL-3 samples. Though our primary focus is on infectious disease and immune response, our services are available to investigators in all fields. To learn more about our services, follow these links or feel free to contact us if you have any questions.
Flow cytometry basics
Flow cytometry is a method of looking at cells or other suitable particles using fluorescence and qualities of light scattering. There are several excellent introductions to flow cytometry and cell sorting readily available on the internet. Here are just a few of the most useful sites.
- Introduction to flow cytometry — From ABD Serotec. A concise and informative overview of flow cytometry and cell sorting principles and practices. Basic information everyone should be familiar with before embarking upon a flow cytometry experiment.
- Compensation: an informal perspective — From Mario Roederer. One of the most respected leaders in the field of flow cytometry provides a detailed look at fluorescence compensation and how it can make or break your multi-color assay results.
- Practical Flow Cytometry, 4th edition — by Howard Shapiro. Far from “just the basics”, this is THE textbook for all things flow cytometry. The fourth edition is kindly available online for free and can also easily be accessed from your mobile device.
- Fluorescence spectrum viewer – From BD. This web-based interactive tool can assist you in your search for the right combination of colors for your experiment and show the amount of spillover between fluorochromes.
- Table of fluorochromes – From the Salk Institute. Basic but useful information about a wide variety of fluorescent dyes and proteins.
Cytometry and sorting policies
If your research is done according to an MU Institutional Biosafety Committee (IBC) protocol, an amendment must be submitted to add the LIDR Immunology Core to your protocol. If we have not worked with your organism before, the IC will have to add it to our own protocol as well. Please allow at least a couple of weeks for this process.
All samples must be filtered with a 40 micron or smaller cell strainer immediately prior to bringing the samples to the IC, even if filtering is an earlier step in your cell preparation protocol. Filters can be purchased from Fisher Scientific as catalog number 22-363-547.
If your samples are fixed BSL-3 agents, you must validate your fixation protocol and submit it to the biosafety officer.
To make an appointment for analysis, or sorting, contact the core technical manager. If any antibodies, fluorescent conjugants, cell-types or infection states have changed since the last experiment, make sure to let her know in your email. Even if all aspects of your experiment are the same, you must fill out and bring a cytometry information sheet and a sorting information sheet (if applicable) to each appointment.
Reminder: All samples must be filtered with a 40 micron or smaller cell strainer immediately prior to bringing the samples to the IC, even if filtering is an earlier step in your cell preparation protocol.
Fluorescence microscopy policies
Contact us for more information.
Flow cytometric analysis (operator-assisted only):
- University of Missouri research — $85.00 per hour
- Other academic or industry users — contact us
Cell sorting (operator-assisted only):
- University of Missouri research – $150.00 plus $100.00 per hour
- Other academic or industry users — contact us
- There are no fees for use of the microscope at this time.
Flow cytometry and cell sorting
All analysis and sorting is performed on the Beckman Coulter MoFlo XDP high speed cell sorter — a powerful three-laser, 10-parameter (FSC, SSC and 8 colors) cytometer — by a highly trained operator.
- Three fixed-wavelength lasers: 488nm, 561nm and 635nm — ideal for working with green and red fluorescent proteins as well as a variety of dyes.
- Up to 4-way bulk sorting as well as single-cell sorting into 96-well plates or onto microscope slides.
- Analysis speeds up to 100,000 events per second and sorting speeds up to 70,000 events per second (actual recommended speeds vary according to cell size, type and conditions).
- An open platform that allows a high degree of customizing for specific fluorochrome combinations.
The Beckman Coulter MoFlo XDP high speed cell sorter
The chart below illustrates the excitation and detection wavelengths available on the MoFlo XDP along with common fluorochromes that can be used. Other fluorochrome options may be available.
Nikon Eclipse 50i trinocular fluorescence microscope – Live BSL-2 and freshly fixed BSL-3 samples can be observed and imaged within the core.
- 10X, 40X, and 60X Plan Fluor objectives
- Filter cubes for DAPI, FITC, TexasRed, and Cy5
- Q-Imaging Systems QiCam Monochrome camera for high resolution fluorescence imaging
- Q-Imaging Systems imaging software
Cytometry information forms
This form must accompany each set of samples brought to the IC for analysis or sorting. If you have any questions about the information requested, contact us.
Sorting information forms
This form, along with a Cytometry Information Form, must accompany each set of samples brought to the IC for sorting. If you have any questions about the information requested, contact us.