You do use the same protocol for SF9 and HI-5 cells.

The basic idea is that you let the cells grow to a desired density (usually between 1 and 2 million per mL), spin them down gently, resuspend them in your freezing media (we just use fresh media with BSA and DMSO), aliquot them in 2ml cryo tubes, and leave them at -20 for 1 hour, then at -80 for roughly 12-18 hours, and then store them long-term in liquid nitrogen. The shelf-life if frozen this way is multiple years, and it's also a great way to store your baculovirus-infected cells which increases the amount of infections you can perform with V1 viruses.

The recipe for freezing media we use is as follows: (Per litre, make 2 falcon tubes)

Take 45ml of fresh, serum-free media and add to a falcon tube

Add 0.5 g of BSA and fully dissolve. Vortex if necessary.

Once dissolved, filter through a sterile 0.22um syringe filter

Add 5ml of *STERILE AND AUTOCLAVED* DMSO to the media

Use this to resuspend your cells. Make the media under a laminar flow cabinet and be absolutely sure to keep your media sterile.

Michael Adams Much tnx for your response but actually usually we use FBS instead BSA in our lab.Actually i do not use when we exactly use BSA instead FBS,because as you know FBS provides some essential nutrients or the cells.could you please tell me why you use BSA? and other question is that i've seen in so many protocol that they do not use media for freezing cells and again whats the use of using it.Im really thankful again for your response.

REGARDS

Hez Majid, we use serum-free growth media especially made for insect cells so that there is no batch-to-batch variability, as there can be with FBS. We add the BSA because we don't use FBS, and need some stabilising agent in the media.