Defined and Scalable Generation of Hepatocyte-like Cells from Human Pluripotent Stem Cells

The overall goal of this protocol is to produce human hepatocyte-like cells from pluripotent stem cells using recombinant laminin matrices under defined conditions.This method allows the user to custom-make functional human hepatocytes to study liver biology in the dish.The main advantage of this technique is that its defined nature allows mass production of hepatocytes with high degree of experimental reproducibility.To begin the experiment thaw 100 microgram per milliliter stock of recombinant laminin-521, or LN-521, at four degrees Celsius.Dilute the thawed LN-521 in ice cold 1xDBPS with calcium magnesium to make a five microgram per milliliter solution.Add one milliliter of the five microgram per milliliter LN-521 solution to coat one well of a six well plate, and rock the plate to spread it evenly in the well.Next, incubate the plates in a 37 degree Celsius 5%CO2 cell culture incubator for at least two hours for urgent use.Take the coated plate from the incubator to the tissue culture hood.Carefully aspirate the coating LN-521 solution without damaging the coated surface.Then immediately add one milliliter of pre-warmed mTeSR