Early in cell culture, heat inactivation was considered necessary to destroy heat-labile complement proteins. Triglia and Linscott 1 determined that levels of complement components in commercial fetal bovine serum FBS were a fraction of adult levels and in 10 samples of commercial FBS, no hemolysis was detected even in undiluted serum.
Serum used to be filtered through 0. There was valid concern that adventitious agents such as mycoplasma could still be present after filtration. In such cases, heat inactivation could help to reduce the potential for obtaining contamination through this route. Today, serum is typically filtered through 0.
This has essentially eliminated the need to heat inactivate serum due to the concern over contamination from serum. Although there are few published papers concerning heat inactivation of serum and its effect on cell culture systems, a report by Hyclone 2 and our own experience at the TCF suggests that in most cases heat inactivation is not required and is, in many cases, detrimental to the growth promoting capacity of serum.
When one heat inactivates serum, not only is complement protein degraded, which has been indicated above may not be necessary , but also all other components such as amino acids, vitamins, growth factors, etc. Allow serum to thaw completely and to reach the temperature of the environment.
Overnight exposure is usually adequate. Remove serum and allow to reach room temperature prior to use. Serum should never be taken from a frozen or refrigerated state and placed in a water bath to expedite thawing or warming.
Such handling compromises the quality of the serum and may cause the bottle to break. However, the practice is labor-intensive and expensive. The protocol must be followed exactly as too high a temperature or too long a time may destroy some growth factors.
On occasion heat inactivation of serum is used because of previous history in the laboratory or for the convenience of stocking only one kind of serum. In past years, Coriell used all heat-inactivated serum for its cell cultures to inactivate the complement protein found in newborn calf serum.
Since we have changed to fetal bovine serum, we find that heat inactivation is not necessary for most cell lines.
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