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. Author manuscript; available in PMC: 2020 Nov 18.
Published in final edited form as: ACS Chem Biol. 2011 Sep 29;6(11):1287–1295. doi: 10.1021/cb200238k

Figure 5. Cultured CNS neurons from w1118 and 4-4, but not those from 2d/2d, incorporate sialo-glycoconjugates in the plasma membrane that were sensitive to sialidase treatment.

Figure 5

Cells were cultured for 2-6 days in serum-containing medium and 250μM NAZ (or NAC) (1-3 CNS/coverslip). The Click-iT reaction with Alexa-Fluor-488-Alkyne was carried out in live cells; while the immunostaining with anti-HRP and nuclear labeling with Hoechst were done following fixation. (A-B) Pictures of neurons taken using the Apotome Zeiss system, (63x objective, bar=10μm), panels going from left to right show pictures of 1μm thickness taken from the bottom towards the top. (C) Mean percentage of neurons that incorporated Sia5NAz in the plasma membrane. Sia5NAz incorporation was measured in 62-121 neurons per condition in each experiment. (n=# experiments). One-way analysis of variance (P<0.0001); post test Bonferroni, $P<0.001 between the group and the 2d/2d group. No significance difference was found between the w1118 and the 4-4 groups. (D-E) Pictures taken with the Apotome Zeiss system, (1μm thickness, 63x objective, bar=10μm) of untreated and sialidase treated cells. (F) Mean percentage of w1118 and 4-4 neurons that incorporated Sia5NAz in the plasma membrane in untreated and sialidase treated cultures. Sia5NAz incorporation was measured in 78-121 neurons per condition in each experiment. n=3 separate experiments. One-way analysis of variance (P<0.0001); post test Bonferroni, $P<0.001 between the sialidase treated and untreated group. No significance difference was found between the untreated w1118 and the 4- 4 groups.