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After completing all sulfation/desulfation steps, the sample with only a single desulfation results in a fairly uniform sulfur distribution along the z-axis inside of the monolith.
For all the purification steps, the sample fractions were assayed for protein content and protease activity using casein as substrate.
By omitting the elution steps, the sample handling time is shortened and sample losses are less likely to occur (Pinto et al. 2010).
After the washing steps, the sample was resuspended in 500 μl BSA buffer and was ready for flow cytometry (Becton Dickinson, Mountain View, CA, USA).
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The particle filter consists of three sequential steps: the sampling, the importance factor, and the resampling step.
After completing all of the fabrication steps, the samples are annealed in vacuum to improve the device characteristics.
After the corresponding incubations and PBS washing steps, the samples were finally mounted with Mowiol (Calbiochem, Merck Chemicals) following standard protocols [31].
During all preparation steps, the samples were kept under cooled conditions.
After three washing steps the samples were resuspended in 250 μl PBS.
For BrdU staining, after the described steps, the samples were fixed in 4% PFA in PBS for 20 min at RT.
After washing steps, the samples were briefly incubated in chemiluminescence reagent Plus (Perkin Elmer) and the bands were detected using Kodak Image Station.
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