Expression of mfap4, mpeg1, ptpn6, and cxcr3.2 is dependent on Spi1 transcription factor. (A-L) Whole-mount in situ hybridization. Wild-type expression of genes mfap4 (A,C), mpeg1 (E,G), and ptpn6 (I,K) as shown in whole embryos and enlarged tail regions shows a pattern typical for myeloid cells. Expression of genes mfap4 (B,D), mpeg1 (F,H), and ptpn6 (J,L) is absent in spi1 knockdown embryos. Images were taken with a Leica M165C stereomicroscope equipped with DFC420 camera. Composite images of different focal planes were generated using Adobe Photoshop. (M-P) Single fluorescent in situ hybridization using TSA Cy3 Plus detection. Wild-type expression of gene cxcr3.2, as shown in the tail region (M) and in the magnified region of the blood island (O), shows a pattern typical for myeloid cells. Expression of gene cxcr3.2 (N,P) is absent in spi1 knockdown embryos. Images were taken with Leica TCS SPE confocal microscope, using the 532-nm laser line for scanning of signal in the red channel. The HC PL Fluotar 10.0×/0.30 dry objective was used. The images were processed in ImageJ Version 1.43 software (National Institute of Mental Health). Bars represent 100 μm (M-N) and 60 μm (O-P).