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. Author manuscript; available in PMC: 2017 Jun 1.
Published in final edited form as: Expert Rev Proteomics. 2016 Jun;13(6):609–626. doi: 10.1080/14789450.2016.1190651

Figure 4. Proof of principle for developing a urine-based glycosylated hemoglobin (A1c) assay.

Figure 4

Different dyes were incorporated into hydrogel nanoparticles to determine which nanoparticle bait (dye) was optimal for harvesting A1c. 100ng A1c (LEE Biosolutions cat#325-10, 15kD) was added to 5mL of Surine™ (synthetic urine negative control, Sigma-Aldrich) prior to adding the nanoparticles. Following elution from the nanoparticles, western blotting with anti-A1c (LifespanBio, LS-C194149-250) was used to demonstrate A1c capture by the nanoparticles. Reactive Black 31 (Lane 7) was found to be the optimal dye for harvesting A1c. Lane 1: BenchMark pre-stained protein ladder, Lane 2: Magic Mark chemiluminescent protein standard, Lane 3: 50ng A1c, Lane 4: Trypan Blue, Lane 5: Congo Red, Lane 6: Direct Blue 2, Lane 7: Reactive Black 31, Lane 8: Reactive Blue 221, Lane 9: Remazol Brilliant Blue, Lane 10: Sequential harvesting with Trypan Blue then Reactive Blue 221 nanoparticles, Lane 11: Surine™ sans nanoparticles, Lane 12: empty. (Lanes 4-10=proteins eluted from the respective nanoparticle).