Materials and Reagents

1. Pipette tips
2. Sarstedt 96-well plate (Sarstedt, catalog number: 83.3924 ) (or similar)
3. 1.5 ml tubes
4. Purified virus stock (1 μg of virus per measured well; stored at -80 °C)
   Purification of enteroviruses can be done using either 5-20% or 10-40% sucrose gradient and is described in detail in the original publication (Ruokolainen et al., 2019). Also, CsCl purification may be used but it was observed by us to have more variation from batch-to-batch than sucrose purified virus. The protocol has been tested and observed to work with echovirus 1 and coxsackieviruses A9 and B3 suggesting its wide applicability for enteroviruses and probably for picornaviruses in general.
5. Buffer/solution of your interest
   In our paper we used a wide spectrum of concentrations of different ions and albumin to study the virus priming and opening. As an example, DPBS solution supplemented with 0.01% BSA resulted in high amount of intermediate virus particles.
6. SYBR Green II RNA gel stain (Invitrogen; ThermoFisher Scientific, catalog number S7564 ; stored at -20 °C)
7. RNase A, 10 mg/ml (ThermoFisher Scientific catalog number EN0531 ; stored at -20 °C)
8. 150 mM NaCl solution
9. Ice
   

Equipment

1. Pipette with a volume range including 100 µl
2. 8-channel multipipette with a volume range including 50 µl, or similar (optional)
3. Perkin Elmer 2030 Multilabel Reader Victor X4 (or similar fluorescence plate reader with suitable filter options)
   

Software

1. Perkin Elmer 2030 Manager
2. Microsoft Excel
   

Procedure