The invention relates to a hemispherical
microarray substrate for three-dimensional
cell sphere culture and a preparation method of the hemispherical
microarray substrate. The surface of a
cell culture substrate disclosed by the invention has a regular hemispherical
microarray structure. The method comprises the following steps: pasting a surface
adhesive on a plane, pasting a screen on the plane, fixing microspheres in micropores of the screen, and uncovering the screen to obtain the plane with a
microsphere array;
coating a light-transmitting substrate with
ultraviolet glue, pressing the plane with the
microsphere array on the
ultraviolet glue, performing curing, and separating the light-transmitting substrate from the plane with the
microsphere array to obtain a semi-microsphere array template; and adding an
agarose solution, performing curing, and separating cured
agarose from the microsphere array template to obtain the hemispherical micropore array substrate for three-dimensional
cell sphere culture. The hemispherical micropore array structure provided by the method is formed by simple hemispherical array
rollover, and is easy to operate, low in cost and free of any complex operation. The
cell culture substrate disclosed by the invention can be used for conveniently and massively preparing cell balls with uniform sizes.