Figure 5.

Hemocyte types are separated with Percoll gradients and adoptively transferred

(A) Hemolymph from donor crayfish containing EdU-labeled hemocytes was separated into two layers (1, 2) with Percoll gradients (see Methods for details).

(B) Layer 1 contains hyaline (filled arrowheads) and semigranular (open arrowheads) hemocytes. The hyaline cells have a narrow, clear rim of cytoplasm surrounding a large nucleus. Semigranular cells contain sparse granules and are often spindle-shaped in these preparations.

(C) Layer 2 contained granular hemocytes (arrows), which are spherical cells densely packed with granules.

(D) When hemocytes from layer 2 (granular cells, arrows) were injected into recipient crayfish, these were often found near, but never in, the niche of recipients. Characteristically, the EdU labeling observed was cytoplasmic, suggesting that EdU-labeled donor cells took up EdU during phagocytosis of debris, or perhaps by phagocytosis of other labeled hemocytes.

(E) When cells from layer 1 (hyaline and semigranular cells) were injected into recipient crayfish, cells with EdU nuclear labeling were observed in the niche (arrows) and were similar in size to the intrinsic niche cells. Scale bars: (B, C) 10 μm; (D, E) 50 μm.