Hcy reagent

The blood sample was collected from the antecubital vein after an overnight fasting in each participant. The blood sample was then separated through centrifugation at 3,000 g for 5 min. The level of Hcy was detected by enzymatic cycling assay using Hcy reagent (Beijing Strong Biotechnologies, Inc, Beijing, China) and the ROCHE Cobas 8000 automatic biochemical analyzer (Roche Ltd., Switzerland).

Male Sprague-Dawley rats (12 weeks old) were purchased from Sino-British SIPPR/BK Laboratory Animal Ltd. (Shanghai, China) in the whole study. All of the procedures were guided and approved by the Animal Care and Use Committee of the Second Military Medical University and conducted specifically to the principles of the Institutional Animal Care.
Animals were divided into two groups. The HCY group at the age of 12 weeks was given by L-methionine (1 g/kg body weight per day) in tap water for a period of 8 weeks, as previously described [18 (link)]. The amount of water drank by animals every day was divided into two parts. The first part was mixed with L-methionine according to body weight of the individual, while the second one was the normal potable water without L-methionine. The control group was given equal volume of water. Eight weeks after HCY treatment, plasma HCY levels in rats were measured. Blood samples were taken from rats and were centrifuged for 10 min at 1500 rpm at 4°C, followed by collection of plasma for the evaluation of HCY. HCY reagent (Beijing Strong Biotechnologies Inc., Beijing, China) was used to detect the levels of HCY by an enzymatic cycling method with Beckman AU 5800 automatic biochemical analyzer (Beckman, USA).