Lipid supplements rich in PUFAs such as flaxseed are susceptible to biohydrogenation in the rumen. To prevent ruminal biohydrogenation, various methods have been tested with limited success. In this study, a novel method to “rumen-protect” flaxseed is proposed, which encapsulates flaxseed using a proprietary method (12BT40; N3Feed LLC; Tualatin, OR). To determine whether 12BT40 decreases rumen biohydrogenation of dietary PUFAs, we used a double 3 × 3 Latin square design; 6 mid- to late-lactation, pregnant Holstein cows (1 block each for primaparous and multiparous cows) were fed 0 kg/d (Negative Control), 3 kg/d of 12BT40 (Treatment), and 3 kg/d of the unprocessed ingredients of 12BT40 (Treatment Control) as top-dressing for 2-week periods each. Milk and serum samples were collected at the end of each 2-week treatment period and analyzed for their fatty acid profile and metabolic indicators, respectively. Individual feed intake and milk yield and components were measured to evaluate the effect of 12BT40 on production parameters. Data were analyzed using PROC MIXED in SAS version 9.4. Fixed effects were treatment, period, and parity. Repeated measures within cows were modeled with the random statement. Compared with Treatment Control, 12BT40 increased the proportion of omega-3 in milk FAs from 1.49 ± 0.23wt% to 2.32 ± 0.23% (P = 0.005) (Negative control: 1.07 ± 0.23%). 12BT40 attenuated the flaxseed-associated increase in trans MUFAs (wt%) from 8.23 ± 0.78% to 5.51 ± 0.78% (P = 0.005) (Negative control: 2.52 ± 0.78%). For production characteristics and metabolic serum indicators, 12BT40 and its unprocessed ingredients did not significantly differ, with the exception that 12BT40 attenuated the flaxseed-associated increase in DMI from 23.3 ± 0.7 to 22.0 ± 0.7kg/d (P = 0.04) (Negative control: 20.6 ± 0.2kg/d) and increased milk lactose content from 4.84 ± 0.06% to 4.91 ± 0.0.06% (P = 0.007) (Negative control: 4.80 ± 0.06%). Thus, we conclude that 12BT40 increases omega-3 content of milk by protecting dietary omega-3 in flaxseed from ruminal biohydrogenation.