24
TUNEO
YAMADA
Fig. 6 and Table III it is clear that on addition of ammonium sulphate,
the mesoderm inducing ability carried by fraction AP passes to the
precipitate (AMPt), and not to the supernate (AMSp). Further it is
Guinea-pig bone marrow
ι
homogenize in 0-25 Μ sucrose containing CaCl 2
centrifuge at 2,000 χ g for 5 min.
Supernatant
centrifuge at 100,000 xg for 90 min.
Supernatant
bring to pH 4-8 by adding N/5 acetic acid
Precipitate
I
dissolve in Tris buffer (pH 7-4)
precipitate with 0-5 saturated (NH 4 ) 2 S0 4
Precipitate
I
dissolve in Tris buffer
I
re-precipitate with 0·5 saturated (NH 4 ) 2 S0 4
dissolve in Tris buffer
Supernatant
centrifuge at 100,000 χ grfor 150 min.
Supernatant
Sediment
precipitate with (NH 4 ) 2 S0 4
Precipitate
dissolve in Tris buffer
disperse in
Tris buffer
dialyze against Tris buffer
remove insoluble materials by centrifligation
AP
AMPt
UISp
UlPt
AMSp
FIG. 6. Scheme for fractionation of bone marrow.
definitely shown that after ultracentrifugation the mesoderm inducing
ability is localized in the supernate (UISp). Now according to sedimentation diagrams prepared by K. Takata, fraction AMPt is composed
of a major component sedimenting slowly and a minor one sedimenting
relatively quickly. After ultracentrifugation the main component