Whole blood and RBC aliquots must be deproteinated before injection into the chemiluminescence reaction vessel to avoid excessive foaming. If processing previously frozen samples, they should slowly thaw on ice. Proteins are precipitated by adding cold methanol to samples in a 1:1 ratio (v/v) and mixture vortexing. Proteins are then removed from mixture by centrifugation of the mixture at 13,000×g for 5 min at 4°C. Supernatant is carefully transferred to a new 1.5 mL tube and used for nitrite analysis (seeNote 13 and 14 ).
It is necessary to keep small aliquots (~1 mL) of all solutions used in your sample treatments––stop solution, methanol, water, or any additional treatment that was used. All these solutions might contain nitrite contamination, which will contribute to the nitrite concentration measured in samples. Nitrite content in all these solutions is determined at the same time as nitrite in samples and, if necessary, corrections for nitrite contamination from these external sources can be made.
It is necessary to keep small aliquots (~1 mL) of all solutions used in your sample treatments––stop solution, methanol, water, or any additional treatment that was used. All these solutions might contain nitrite contamination, which will contribute to the nitrite concentration measured in samples. Nitrite content in all these solutions is determined at the same time as nitrite in samples and, if necessary, corrections for nitrite contamination from these external sources can be made.
