1. What kind of beads do I need to use?
The accuracy of the software is very much dependent on the beads. They therefore need to be of known diameter and refractive index. A number of companies sell 'NIST-Traceable' particles that have had their diameter verified by electron microscopy and their refractive index reported. These type of beads will offer optimal accuracy.
2. How many beads do I need to use?
It is recommended at least 10 bead populations are used.
3 Why does the parameter I want to convert not appear in the dropdown menu?
When files are imported into the software, only the common parameters across all of the files are listed. This is to make sure that when attempt to convert a parameter every file can be converted. If a parameter is missing from just one of the files, or its name appears differently in a file, that parameter will not be listed in the dropdown menu. If this happens, put the files that have common parameter names into a folder and import this folder.
4. Why does it sometimes take a long time to calculate?
The FCMPASS software works using a database. This database is created for every unique refractive index inputted into the software. If you calculate a model with a refractive index that has already been calculated the software can simply look up the data. If, however, you use a new refractive index for a bead input (this may be due to using an illumination wavelength not pre-calculated) or change the variables of the default extracellular vesicle models then a database will have to be created for these specifications. The next time this specification is used, however, the calculation should occur much faster as the database will already exist.
5. Can I reimport the exported fcs files into my Flow Cytometer's software?
While the exported fcs files may be compatible with a Flow Cytometer's native software, they are only guaranteed to work with third-party analysis softwares such as FlowJo and FCSExpress. This is because third-party analysis softwares are far less restrictive on the type of files that can be imported.