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A case of e-cigarette-induced polycythaemia (increased haemoglobin and red cell numbers) of a 70 yr man who had smoked about 25 cigarettes per day for 50 years until he was 70 years old and switched from conventional smoking to e-cigarettes (electronic nonnicotine delivery system, approximately 40 puffs per day) one year before being investigated

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Immunohistochemistry was performed as follows: free-floating VTA brain sections were incubated 1 h at 4 C in a blocking solution of phosphate-buffered saline (PBS) containing 3% bovine serum albumin (BSA, Sigma A4503) and 0.2% Triton X-100 and then incubated overnight at 4 C with a mouse anti-tyrosine hydroxylase antibody (TH, Sigma, T1299) at 1:500 dilution in PBS containing 1.5% BSA and 0.2% Triton X-100 (see supplementary Table 1)

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Again, nicotine patches to support abstinence were offered to participants after the third session