4.1 Cell source and culture conditions of LAD2 cell line
LAD2 cells are often sourced from the Laboratory of Allergic Diseases,
National Institutes of Health (NIH) in Bethesda, United States of
America (USA) where the cells were first discovered and successfully
cultured. Of the 75 papers analysed, 55 of the research groups obtained
their LAD2 cells from NIH, specifically the laboratory of Drs Arnold
Kirshenbaum and Dean Metcalfe 47. Some were obtained
from Otwo Biotech company (1 of 75), Dr Yangyang Yu of Shenzhen
University (1 of 75), Dr Michael of Colombia University and Professor
Renshan Sun of Third Military Medical University (1 of 75). However, the
authors did not report the original source of their gifted LAD2 cell
line. Finally, 17 research groups did not specify the origins of their
LAD2 cells. Figure 3 summarizes the most common sources to obtain LAD2
cells.
StemPro-34 is the culture medium used to culture LAD2 cells as observed
in most of the 75 papers analysed (74 of 75). There is only one
publication that reported the use of IMDM with supplementation of 10%
FCS to culture the LAD2 cells. StemPro-34 is a specifically formulated
serum-free medium used to support the growth of human hematopoietic
cells. Supplements are often added in the StemPro-34 culture medium for
LAD2 cells. The medium is usually supplemented with StemPro-34 nutrient
supplement which include pre-mixed penicillin/streptomycin, L-glutamine,
and human stem cell factor. Apart from that, Zou et al. (2022) added 50
ng/mL of IL-6 into their StemPro-34 medium. Cytokines and growth factors
are added into the medium to further support the growth of the
progenitor cells. The use of StemPro-34 is in accordance with the medium
used when LAD2 cells were first cultured in the laboratory of Drs
Kirshenbaum, Akin and Metcalfe. Similarly, the medium was supplemented
with 2 mM L-glutamine, 100 IU/mL penicillin, 50 µg/mL streptomycin, 100
ng/mL rhSCF. The addition of 100 ng/mL of rhIL-6 and 30 ng/mL rhIL-3 for
the first week were optional 45. Kirshenbaum et al.
(2003) also noted that even though LAD2 cells could survive in medium
without SCF, the numbers doubled in approximately 3 weeks with 100 ng/mL
SCF, and the addition of rhIL-3, rhIL-5 or rhIL-6 did not influence the
cell numbers. SCF is needed for the growth of LAD2 cells as they do not
possess the c-kit activating mutation at codon 816. LAD2 cells
were grown in a humidified incubator at 37°C with 5%
CO2. Table 3 summarized the culture medium used to grow
LAD2 cells. As suggested by Rådinger et al. (2010), hemidepletions of
the medium were performed by most researchers when growing the LAD2
cells at the frequency of once a week. The cell density was generally
maintained between 1 × 105 to 2 ×
106 cells/mL.