Techniques to accelerate the availability of human keratinocyte grafts

Human keratinocytes grown in cultures have been used successfully in the coverage of large burns. The time required to grow sufficient human keratinocytes for grafting is a major limitation of this technique. Three variables--plating density, length of time to confluence in secondary culture, and media supplementation--were studied to find conditions that increase the rate of keratinocyte growth. Normal human keratinocytes were used for all studies. Plating densities varied between 2.5 x 10(3) and 3.7 x 10(4) cells per square centimeter. Time in secondary culture was studied daily from day 2 to day 17. Media supplements included fetal bovine serum, human serum, and three commercially available serum supplements. These studies indicate that human keratinocytes, grown for wound grafting, can grow more rapidly to a higher cell number if they are plated at a density of 2 x 10(4) cells per square centimeter, passaged by 8 to 10 days or before confluence slows cell growth, and grown in 5% fetal bovine serum, 1.25% fetal bovine serum, or 5% human serum. Use of these techniques may decrease the time necessary for wound coverage with human keratinocytes.