Isolation and molecular characterization of hemocyte sub-populations in kuruma shrimp Marsupenaeus japonicus
Crustacean hemocytes, which have usually been classified morphologically based on methods using Giemsa or May-Giemsa stains, have recently been categorized using monoclonal antibodies or marker genes. However, these latter techniques are not yet widely used, and different classification methods are used for hemocytes among laboratories. Therefore, we aimed to develop a molecular classification method that can be widely used by researchers. The method we have developed uses lectins and magnetic-activated cell sorting (MACS) to isolate sub-populations of hemocytes. Two lectins, wheat germ agglutinin (WGA) and tomato lectin (Lycopersicon esculentum lectin; LEL), characteristically bind to hemocytes, which allows them to be classified into three sub-populations. Furthermore, different sub-populations of hemocyte can be isolated by using LEL and MACS. These sub-populations were characterized as non-granular and granular hemocytes, and the accumulation patterns of the gene transcripts were consistent with the results of a functional analysis reported previously. The lectin-based hemocyte isolation method developed in this study has good reproducibility.
KeywordsLectin staining Transcriptomics Magnetic-activated cell sorting Crustacean Invertebrate Immunity
This work was supported by the Japan Society for the Promotion of Science (JSPS) Grant-in-Aid for a JSPS research fellow (grant no. 16J08185) to K. Koiwai and a Grant-in-Aid for Scientific Research (A) (grant no. 15H02462) to I. Hirono. All the sequences from total, LELDim and LELStrong hemocytes are archived together with the raw data at the DDBJ Sequence Read Archive under accession no. DRA007926.
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Conflict of interest
The authors declare that they have no conflict of interest with regard to the contents of this article.
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