The freezing characteristics of genetically modified lymphocytes obtained f
rom a donor with mucopolysaccharidosis type II (MPS II) were determined usi
ng cryomicroscopy and controlled rate freezing studies to determine posttha
w viability. The cells from a donor with MPS II used in this investigation
were cultured and transduced with a retroviral vector fbr the iduronate-2-s
ulfatase (IDS) enzyme for clinical studies for human gene therapy. The wate
r transport and intracellular ice formation (IIF) characteristics of the ce
lls were determined after completion of the culture and transduction protoc
ol. The water transport parameters, L-pg and E-lp, for the cultured and tra
nsduced cells were determined to be 4.4 +/- 1.3 x 10(-14) m(3)/Ns and 173 /- 25 kJ/mol, respectively. The IIF nucleation parameters, kappa and Omega,
were 5.5 x 10(10) K-5 and 3.5 x 10(11) (1/m(2) s), respectively. The postt
haw viability of the genetically modified cells was less than the viability
of the freshly isolated cells from the same donor. The postthaw viability
of the cultured and transduced cells from a donor with MPS II was also less
than that observed with cells from a normal donor that were frozen and tha
wed under the same conditions. These studies are essential in understanding
the biophysical changes resulting from the ex vivo culture of cells and th
e manner in which these changes influence the ability of the cells to be cr
yopreserved.