Larval Lymph Gland Dissection: Isolating the Drosophila Hemocyte-Producing Organ
Published: April 30, 2023
Abstract
Source: Reimels, T. A. and Pfleger, C. M. Methods to Examine the Lymph Gland and Hemocytes in Drosophila Larvae. J. Vis. Exp. (2016).
This video describes the Drosophila lymph gland—the primary site of hematopoiesis in the fly larva—and shows an example protocol to dissect and mount the organ for immunohistochemistry and fluorescent imaging.
Protocol
This protocol is an excerpt from Reimels and Pfleger, Methods to Examine the Lymph Gland and Hemocytes in Drosophila Larvae, J. Vis. Exp. (2016). 1. Larval Lymph Gland Immunohistochemistry NOTE: The lymph gland is located approximately one-third length from the anterior end of a larva slightly below the brain on the dorsal side. (See arrow in Figure 1B.) The lymph gland…
Representative Results
Figure 1: In Vivo Crystal Cell Melanization. A) Larvae were placed individually in PCR tubes prior to heating. Red arrows indicate larvae that are not at the bottom of the tubes. B) A typical crystal cell melanization pattern after heating, which sometimes reveals the lymph gland (arrow; dorsal side shown, anterior is left). Scale bar represents 1 mm. <a href="https://www.jo…
Materials
| PBS tablets | MP Biomedicals | 2810305 | |
| dissecting dish | Corning | 7220-85 | |
| microcentrifuge tube | Denville | C2170 | |
| silicone dissecting pad, made from Sylgard 184 kit | Krayden (distributed through Fisher) | NC9644388 (Fisher catalog number) | Made in petri dish by mixing components of Sylgard elastomer kit according to manufacturer instructions. |
| stereomicroscope | Morrell Instruments (Nikon distributor) | mna42000, mma36300 | Nikon models SMZ1000 and SMZ645 |
| tissue wipe | VWR | 82003-820 | |
| forceps | Electron Microscopy Sciences | 72700-DZ | |
| p200 pipette | Eppendorf | 3120000054 | |
| formaldehyde | Fisher | BP531-500 | |
| Triton | Fisher | BP151-500 | |
| Tween 20 | Fisher | BP337-500 | |
| bovine serum albumin | Rocky Mountain Biologicals | BSA-BSH-01K | |
| normal goat serum | Sigma | G9023-10ML | |
| normal donkey serum | Sigma | D9663-10ML | |
| 200 proof ethanol | VWR | V1001 | |
| N-propyl gallate | MP Biomedicals | 102747 | |
| glycerol | VWR | EM-4750 | |
| DAPI (4',6-diamidino-2-phenylindole) | Fisher | 62248 | |
| microscope cover glass, 18 mm circular | Fisher | 12-545-100 | |
| glass microscope slides | Fisher | 22-034-980 | |
| 24-well plate | Corning | 351147 | |
| disposable transfer pipet | Fisher | 13-711-9AM | |
| fluorescence microscope | Zeiss | Axio Imager.Z1 |
Citazione di questo articolo
Larval Lymph Gland Dissection: Isolating the Drosophila Hemocyte-Producing Organ. J. Vis. Exp. (Pending Publication), e20117, doi: (2023).