Companies: BioLife Solutions

For thawing cells, conventional practice is to warm the cells quickly in a warm water bath (e.g., 37 C) to just about the point to which the last bit of ice is about to melt and then to dilute the cells slowly into growth media. If the sample is allowed to get to warm the cells may start to metabolize, and be poisoned by the DMSO (dimethyl sulfoxide) that is used in the feezing prcoess. (Schryver, US Patent Application 15,602,711 published as US 2017/0257908 and US 10,917,941, which is a continuation of US Application 14.712,120 published as US 10,555,374)

Thawing Devices and Methods:

Pluristem LTD (CA 2883826) discloses a system for heating a biological material which includes a processor configured to receive an input associated with a target temperature and transmit a signal to controllably move a heating deice relative to the base for a time period, wherein the time period is detemriend based on the target emperature and content volumne. A method for thawing the frozen biological material includes the steps of controllably moving the heating device for a specific time period, wherein the time period is determiend based on the target emperature, the vial content material and the content volumne. Initially, a thawing cycle may active only after heating device has reached an initial temperature. This may reduce the affect of room and/or device temperature on the thawing procedure.

Thawing Protocols for Specific types of Cells: 

Thawing Melanoma cells:

Preparing Media (Dulbeo’s Modification Eagles Medium “DMEM”  ): 

(1) spray under hood with ETOH

(2) Add 50 ml of FBS to our bottel of DMEM. (We have a 500 ml volume and need to use 10% FBS so we will use 50 ml FBS.)

(3) Add 5ml of antibiotic solution (P/S/A “penicillin, streptomycin “antimicotic ) and 5 ml of L-glutamine (since we want a 1:100 dilution)

(4) invert the jugs a couple of times and store at 4c

(5) label “DMEM 10 + P/S/A JL date”

Thawing:

(1) obtain large conical tube (yellow cap) and transfer 30 ml of the media above. Use sterile conditions under the hood.

(2) get cells out from liquid N2 (ex. rack 10, Box 6, B-16; keep rack right on top of machine)

(3) thaw cells quickly in water bath and transfer with 2 ml pipette just as soon as cells have thawed.

(4) mix tube by inverting and then use 25 ml pipette to add the cells in the tube to sterile flask which has been labelled (ex. “B1678H1 F:2/27/94 T: 6/23/03 JLR”)

(5) observe cells under microscope in flask to access survivability  (“alive cells will adhere to bottom of flask and become dendritic. floating cells will be dead by tommorroo)

(6) grow these cells at 37.5 with falsk in horizontal position