Cell-free protein synthesis (CFPS) platforms, once primarily a research tool to produce difficult to express proteins, are increasingly being pursued by the synthetic biology community for applications including biomanufacturing, rapid screening systems, and field-ready sensors. While consistency within individual studies is apparent in the literature, challenges with reproducing results between laboratories, or even between individuals within a laboratory, are discussed openly by practitioners. As the field continues to grow and move toward applications, a quantitative understanding of expected variability for CFPS and the relative contribution of underlying sources will become increasingly important. Here we offer the first quantitative assessment of interlaboratory variability in CFPS. Three laboratories implemented a single CFPS protocol and performed a series of exchanges, both of material and personnel, designed to quantify relative contributions to variability associated with the site, operator, cell extract preparation, and supplemental reagent preparation. We found that materials prepared at each laboratory, exchanged pairwise, and tested at each site resulted in 40.3% coefficient of variation compared to 7.64% for a single operator across days using a single set of materials. Reagent preparations contributed significantly to observed variability; extract preparations, however, surprisingly did not explain any of the observed variability, even when prepared in different laboratories by different operators. Subsequent exchanges showed that both the site and the operator each contributed to observed interlaboratory variability. In addition to providing the first quantitative assessment of interlaboratory variability in CFPS, these results establish a baseline for individual operator variability across days that can be used as an initial benchmark for community-driven standardization efforts. We anticipate that our results will narrow future avenues of investigation to develop best practices that will ultimately drive down interlaboratory variability, accelerating research progress and informing the suitability of CFPS for real-world applications.

Crossbred beef steers (n = 240; 12 pens/treatment; initial BW = 305 ± 17.7 kg) were used in a randomized block design feedlot study to evaluate the influence of coated trenbolone acetate (TBA) and estradiol-17β (E2) implants (Merck Animal Health, Madison, NJ) on gain performance, carcass traits, and sera metabolites. The five treatments were no implant (NI), Revalor-XR on d 0 [200 mg TBA + 20 mg E2 (coated); XR], Revalor-XS on d 0 [200 mg TBA + 40 mg E2 (total): 80 mg TBA + 16 mg E2 (noncoated) and 120 mg TBA + 24 mg E2 (coated); XS], Revalor-200 on d 0 [200 mg TBA + 20 mg E2 (noncoated); E200], or Revalor-200 on d 70 (D200). Interim BW and blood were collected on d 0, 14, 35, 70, 105, 140, and 175 prior to feeding and on d 213 prior to shipping. Following a 24 h clot at 4 °C, sera was harvested to quantify circulating E2, IGF-I, NEFA, serum urea-N (SUN), and 17β-trenbolone (17β-TbOH). Implanted steers had greater (P ≤ 0.05) ADG, G:F, and final BW than NI controls. Implants increased (P < 0.05) HCW by 8%, 366 vs. 391, 414, 380, and 396 ± 6.4 kg, for NI vs. XR, XS, E200, and D200, respectively. The greatest (P ≤ 0.05) dressing percentage, yield grade, and calculated empty body fat occurred in XS, which had greater (P < 0.05) rib fat than NI, XR, and D200. Marbling scores in NI were greater (P < 0.05) than E200 and D200; steers in XR and XS were intermediate (P > 0.10), not differing from NI, E200, or D200. An implant × day interaction (P ≤ 0.01) was noted for circulating E2, IGF-I, SUN, and 17β-TbOH. Implanted steers had elevated (P ≤ 0.05) sera E2, IGF-I, and 17β-TbOH, and decreased (P < 0.05) SUN following implantation compared to NI controls. Serum NEFA differed (P < 0.01) over time, but did not differ (P > 0.10) due to implant treatment. These data indicated that the polymer coating applied to the XR implant delayed release of steroidal hormones congruently to D200, with no negative impact on marbling. The greatest dose of E2, contained in XS, provided improvements in gain and carcass weight without detriment to marbling scores compared to NI.

For more than 60 y, beef cattle producers have safely used various types of growth-enhancing technology (GET) such as steroidal implants with anabolic activity and orally active beta-adrenergic agonists to increase skeletal muscle growth rate, improve carcass leanness, increase average daily gain (ADG), and alter dry matter intake (DMI) compared to non-treated cattle. Generally, the use of a GET increases ADG and only moderately affects DMI relative to non-treated cattle; subsequently, this enhances the rate of live weight gain relative to the amount of feed needed to achieve that gain, this is referred to as feed efficiency (G:F). When a producer chooses to utilize a GET, improvements in treated cattle over non-treated cattle are typically in the range of 8% to 28% for ADG and 5% to 20% for G:F. This review of the literature is intended to provide up to date insight into the mechanisms of how steroidal implants with anabolic activity enhance cattle growth and how these technologies have evolved since their introduction to U.S. beef producers nearly 60 y ago.

No abstract