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8 mM of reducing agent DTT (Sigma) was then added to the samples and incubated in a thermomixer at 30C for 30 minutes, followed by the addition of 20 mM iodoacetamide (Sigma) for protein carboxylation with incubation at 30C in the dark for 30 minutes

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But for new users, seeing pink liquid in a syringe can feel different than seeing clear liquid, even though the technique is identical

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After staining with crystal violet and subsequent destaining, absorbance was recorded at 517 nm using a spectrophotometer, following the protocol described by [19, 23]

Splenocytes from immunized mice were stimulated with 10 g/mL of WS protein for 24 h and in the last 4 h it was added PMA (50 ng/mL), Ionomycin (500 ng/uL), Golgi Plug (BD Bioscience) and Stop Golgi (BD Bioscience)