2-NBDG uptake

2-NBDG uptake assay

The fluorescently labeled glucose analogue, 2-deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]-D-glucose (2-NBDG) serves as a probe for monitoring glucose uptake.

Prepare a 30 mM stock of 2-NBDG (molecular weight: 342 g/mol; Invitrogen N13195) 

• Dissolve 5 mg 2-NBDG in 487 ul ethanol. Store at -20 C in the dark*. 

Perform 2-NBDG staining of larval discs 

• Dissect out the larval heads in 1X PBS (at room temp) on a glass dissection dish. 
• Invert the body cavity such that larval imaginal discs are exposed to the medium. 
• Prepare the working solution of 2-NBDG (final: 300 uM): add 5 ul stock to 500 ul PBS. 
• Replace the PBS with 2-NBDG in PBS. 
• Incubate on a rocking platform or with gentle shaking or rotating in the dark* for 15 min at room temperature. 
• Replace the 2-NBDG in PBS with fresh PBS (for rinsing). 
• Fix the larval tissues with 4% PFA-PBS and proceed according to standard staining protocol. 

*2-NBDG is light-sensitive.

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