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In each of 2 yr a 2 by 2 factorial design of forage type Tifton-85 or Coastal bermudagrass large round bale silage and supplement frequency of daily or 3 d/wk (3X) was used to evaluate growth and reproductive performance in heifers. In each year, heifers (n = 123, BW = 272 ± 3.7 kg) were blocked by breed, BW, and BCS and randomly assigned to 1 of 4 pens (15 heifers/ pen) for a 112 d development period. Heifers were pen fed silage and supplement fed daily or 3X, mean amount was 2.96 kg•heifer-1•d-1. Weekly blood samples were collected for analysis of plasma progesterone concentration. Heifer BW and BCS were collected every 28 d. On d 112, heifers were synchronized using a CIDR and time-AI. Ten d following AI, heifers were exposed to a bull for 60-d. Pregnancy status was determined at 30 and 60 d intervals. Statistical analysis used pen as the experimental unit and PROC MIXED to analyze BW, BCS, and GENMOD for reproductive performance. Fixed effects included forage, frequency, and interactions, random effects included year and heifer. Final BW and 112-d ADG were not different (P > 0.31) for Coastal than Tifton-85 (351 vs 345 ± 7.4 kg and 0.69 vs 0.65 ± 0.11 kg/d). Final BW and 112-d ADG did not differ (P > 0.84) between Daily and 3X supplemented heifers (348 vs 348 ± 4.8 kg and 0.67 vs 0.68 ± 0.12 kg/d). There was no forage x frequency interaction (P = 0.20) for final BW and 112-d ADG. Body condition score on d 112 did not differ (P > 0.05, 5.4 ± 0.11). Cycling status at the initiation of the breeding season tended (P = 0.10) to be greater for Daily (46%) compared to 3X (31%) heifers. Artificial insemination pregnancy exhibited a forage x frequency interaction (P < 0.009). Tifton-85/daily supplemented heifers (73%) had similar (P > 0.05) AI pregnancy rate compared to Coastal/3X (55%), but greater (P < 0.05) than Tifton-85/3X (42%) and Coastal/daily (39%). Coastal/3X and Tifton-85/3X both had greater (P < 0.05) AI pregnancy rate than Coastal/daily. However, 30-d and 60-d pregnancy rates did not differ (P > 0.05) among treatments (73% and 85%, respectively). Similarities in heifer final BW and ADG among forage types and supplement frequency resulted in similar heifer 30-d or 60-d pregnancy rate. However, Tifton-85 did result in greater 30- and 60-d pregnancy rates. [ABSTRACT FROM AUTHOR]

A proof of concept demonstration was designed to evaluate the use of cull, table stock, chip potatoes (var. Atlantic) to upcycle low to marginal quality bahiagrass (Paspalum notatum) hay as an alternative feed resource for beef cattle producers. Two treatments were utilized; treatment one (Trt1) was a combination of 25% hay and 75% potatoes, 3.31 kg of hay and 9.99 kg of potatoes (as-fed basis) were combined; treatment two (Trt2) was a combination of 20% hay and 80% potatoes, 2.5 kg of hay and 9.99 kg of potatoes (as-fed basis) were combined. Two 208-L and one 132-L plastic barrels with lids were utilized for each treatment. Hay was peeled off of the bale and potatoes were rough chopped through a wood chipper. Hay and potatoes (DM = 93.5 and 70.5%, TDN = 61 and 85%, CP = 8 and 10%) were alternately layered into a barrel and compressed. After the sixth layer, barrels were sealed with a matching lid fitted with a check-valve and a vacuum applied to remove air. Barrels were opened and samples taken via a silage core-sampler after 72 d. Two cores from each barrel were collected for analysis. Comparisons of ensilage treatments was conducted with the Mixed procedure of SAS. The statistical model included ensiling treatment and replication. Least squares means are reported. Dry matter content did not differ (P = 0.96, 32 ± 1.4%) between treatments and were 50% below the goal of a 60-65% DM product. Energy values did not differ (P ≥ 0.17) on a DM basis, but were below expectation (TDN = 62.1 ± 2.04%, NEm = 0.26 ± 0.016 mcal/kg, NEg = 0.15 ± 0.015 mcal/kg) based on the proportions used. Crude protein was not different (P = 0.47, 10.1 ± 0.49%) and greater than or equal to predicted. A nearly 50% greater (P = 0.11) amount of starch in Trt2 compared to Trt1 indicated that fermentation of starch was limited. Limited fermentation is indicated by the similar (P = 0.92) lactic acid, but nearly 50% decrease (P = 0.06) in total acid production in Trt2 compared to Trt1. DMI potential, predicted from NDF, was similar (P = 0.27, 13.5 ± 1.28 kg). This conclusion must be taken with caution as no measure of digestibility or intake was established. Proof of concept was successful with this demonstration. Upscaling to a small-scale, on-farm demonstration would seem necessary to validate this conclusion. [ABSTRACT FROM AUTHOR]

Many law enforcement and government agencies have dedicated law enforcement officers (LEOs) who respond to agricultural crime, inspection, urban/rural interface issues, and livestock neglect cases. We developed a training and certification program for LEOs in the field of animal science. Three classes were offered to 52 individuals in a 12-month period. Instruction used a combination of classroom and experiential learning sessions. Daily homework assignments and quizzes were administered to enhance retention. Final assessment to achieve certification included six hands-on exercises to demonstrate proficiency and a written, multiple-choice examination. A total of 94% of the participants passed the certification requirements and overall subject matter knowledge increased by 36%. Likert scale responses (1 to 5 = very little to very much) regarding knowledge of 14 subject matter topics before (2.89 ± 0.11) and after (3.83 ± 0.07) indicated a mean increase of 0.91 ± 0.09 units (UNIVARIATE procedure of SAS, v9.2). Topics with a >1 unit increase included: cattle and equine BCS, equine behavior and learning lab, and nutrition. Our objective was to evaluate the post-course application of the LECALE curriculum. An online survey instrument was developed (Qualtrics™) and distributed to LECALE attendees (n = 50) in June 2016. Twenty-five attendees participated in the survey (50% response rate) providing representation from all three courses (n = 12, 5, 8). Time between completion of the course and administration of the survey was 15, 6, and 3 months, respectively. The majority of attendees agreed that as a result of the topics presented in the LECALE course, they have been and continue to be better prepared when responding to livestock calls (animal identification = 92%, cattle behavior = 96%, equine behavior = 92%, cattle and equine BCS = 94%, euthanasia and disposal = 92%). Attendees indicated they often refer to the information provided during the course regarding horses (66.7%) and cattle (58.3%). Attendees agreed they have been and continue to be better prepared to respond to livestock calls as a result of field trips provided during the course (equine retirement home and beef cattle research unit visits = 91.6%). 33.3% indicated they have utilized the eXtension Horses Body Condition Score app since attending the course. Results indicate that the LECALE program improved the ability of LEOs to perform their job duties. Post-course survey responses are also being utilized to further enhance the LECALE curriculum. [ABSTRACT FROM AUTHOR]

A 2 x 2 factorial design investigated growth, performance, and liver TM status in Angus (n = 16) and Brangus (n = 16) bulls born to cows supplemented with either inorganic (n = 16, 8/breed, Se selenite and salt sulfates) or organic (n = 16, 8/breed, Se-yeast and proteinates) TM sources of Co, Cu, Mn, Se, and Zn. The TM supplement was initiated 82 ± 2 d precalving. At weaning, calves were assigned to 1 of 4 pens based on treatment, receiving the same TM supplement as their dams. All bulls were fed a high energy/protein diet to gain approximately 1.14 kg/d with the TM source delivered in a pellet based on manufacturers recommendations. Bull BW, BCS, and hip height (HH), were collected every 28 d. Liver biopsies for TM status and ultrasound body composition measurements of LM area (LMA), 12th rib fat thickness (FAT), and LM intramuscular fat percentage (IMF) for body composition were collected every 56 d. Data were analyzed as repeated measures using PROC MIXED with bull as experimental unit and fixed effects of TM, breed, time, and appropriate interactions. Results are presented as LSM ± pooled SE. All growth, body composition, and liver TM measurements were affected (P < 0.02) by time. There was no effect (P > 0.05) of TM source on body composition or liver TM concentrations. Inorganic had greater (P = 0.01) mean ADG (1.35 vs. 1.20 ± 0.04 kg/d) but lesser (P = 0.01) mean BCS (4.8 vs. 4.9 ± 0.03) than organic. Pooled across time, Brangus had greater (P < 0.05) LMA (64.95 vs. 60.43 ± 1.46 cm²), HH (119.3 vs. 114.6 ± 0.65 cm), BW (359 vs. 340 ± 6.0 kg), and BCS (4.9 vs. 4.8 ± 0.03) but lesser (P < 0.05) IMF (2.70 vs. 3.60 ± 0.14), than Angus, respectively. Liver Cu and Se were affected by breed x time (P < 0.05). On d 0, breeds were similar (P > 0.05) for Cu (109 ± 24.3 µg/g) and Se (0.94 ± 0.06 µg/g). Whereas, at d 168, liver Cu (297 vs. 148 ± 26.1 µg/g) and Se (1.76 vs. 1.42 ± 0.07 µg/g) were greater (P < 0.05) in Brangus than Angus, respectively. Prenatal and postnatal TM source had no effect on liver TM status of bulls. [ABSTRACT FROM AUTHOR]

A 2 x 2 factorial design evaluated breed (Angus vs. Brangus) and prenatal/postnatal TM source (inorganic vs. organic) effects on bull sexual development. Bulls (241 ± 2 d, 249 ± 4 kg, n = 32, 8 per TM x breed) born to dams supplemented with Co, Cu, Mn, Se, and Zn as inorganic (Se selenite or salt sulfates) or organic (Se-yeast and proteinates) TM sources starting 82 ± 2 d pre-calving. At weaning, calves were assigned to 1 of 4 pens based on treatment, receiving the same TM supplement as their dams. All bulls were fed a high energy/ protein diet to gain approximately 1.14 kg/d with TM source delivered in a pellet based on manufacturers recommendations. Bi-weekly semen collection, scrotal circumference (SC) and BCS (scale 1 to 9) were recorded. Semen collection was initiated when SC was 26 cm. Puberty was defined as an ejaculate with sperm concentrations ≥ 50 x 106 cells/mL, with ≥ 10% motility. Sexual maturity was defined as two consecutive semen collections with SC ≥ 30 cm, sperm motility ≥ 30%, and normal sperm ≥ 70%. Data were analyzed with PROC MIXED with fixed effects of TM source, breed, and interaction for puberty and sexual maturity data. Results are presented as LSM ± SE. At puberty (n = 27), no effect (P > 0.05) of TM source, breed, or interaction for age (337 ± 7 d), BCS (4.8 ± 0.1), motile (33.0 ± 2.3%) or normal sperm (48.7 ± 4.5%) occurred. However, Brangus (172.4 ± 28.2 x 106 cells/mL) had greater (P = 0.05) sperm concentration than Angus (96.9 ± 23.3 x 106 cells/mL). A TM source x breed effect (P = 0.01) occurred for SC at puberty; Brangus inorganic (33.5 ± 0.9 cm) were greater (P < 0.05) than Brangus organic (30.3 ± 0.9 cm), while Angus inorganic (29.9 ± 0.7 cm) and organic (31.2 ± 0.7 cm) were similar (P > 0.05). At sexual maturity (n = 15), organic (351 ± 18 d) were numerically (P = 0.14) younger than inorganic (391 ± 16 d), while BCS (5.2 ± 0.1), SC (33.6 ± 0.7 cm), sperm concentration (156.5 ± 24.9 x 106 cells/mL), motile (47.3 ± 2.7%), and normal (76.4 ± 1.0%) did not differ (P > 0.05) by TM source, breed, or interaction. Pubertal traits were affected by breed but not TM source. However, TM source may decrease bull age at sexual maturity. [ABSTRACT FROM AUTHOR]

Many law enforcement and government agencies have dedicated law enforcement officers (LEOs) who respond to agricultural crime, inspection, urban/rural interface issues, and livestock neglect cases. We developed a training and certification program for LEOs in the field of animal science. Three classes were offered to 52 individuals in a 12-month period. Instruction used a combination of classroom and experiential learning sessions. Daily homework assignments and quizzes were administered to enhance retention. Final assessment to achieve certification included six hands-on exercises to demonstrate proficiency and a written, multiple-choice examination. A total of 94% of the participants passed the certification requirements and overall subject matter knowledge increased by 36%. Likert scale responses (1 to 5 = very little to very much) regarding knowledge of 14 subject matter topics before (2.89 ± 0.11) and after (3.83 ± 0.07) indicated a mean increase of 0.91 ± 0.09 units (UNIVARIATE procedure of SAS, v9.2). Topics with a >1 unit increase included: cattle and equine BCS, equine behavior and learning lab, and nutrition. Our objective was to evaluate the post-course application of the LECALE curriculum. An online survey instrument was developed (QualtricsTM) and distributed to LECALE attendees (n = 50) in June 2016. Twenty-five attendees participated in the survey (50% response rate) providing representation from all three courses (n = 12, 5, 8). Time between completion of the course and administration of the survey was 15, 6, and 3 months, respectively. The majority of attendees agreed that as a result of the topics presented in the LECALE course, they have been and continue to be better prepared when responding to livestock calls (animal identification = 92%, cattle behavior = 96%, equine behavior = 92%, cattle and equine BCS = 94%, euthanasia and disposal = 92%). Attendees indicated they often refer to the information provided during the course regarding horses (66.7%) and cattle (58.3%). Attendees agreed they have been and continue to be better prepared to respond to livestock calls as a result of field trips provided during the course (equine retirement home and beef cattle research unit visits = 91.6%). 33.3% indicated they have utilized the eXtension Horses Body Condition Score app since attending the course. Results indicate that the LECALE program improved the ability of LEOs to perform their job duties. Post-course survey responses are also being utilized to further enhance the LECALE curriculum. [ABSTRACT FROM AUTHOR]

A 2 × 2 factorial design investigated growth, performance, and liver TM status in Angus (n = 16) and Brangus (n = 16) bulls born to cows supplemented with either inorganic (n = 16, 8/breed, Se selenite and salt sulfates) or organic (n = 16, 8/ breed, Se-yeast and proteinates) TM sources of Co, Cu, Mn, Se, and Zn. The TM supplement was initiated 82 ± 2 d precalving. At weaning, calves were assigned to 1 of 4 pens based on treatment, receiving the same TM supplement as their dams. All bulls were fed a high energy/protein diet to gain approximately 1.14 kg/d with the TM source delivered in a pellet based on manufacturers recommendations. Bull BW, BCS, and hip height (HH), were collected every 28 d. Liver biopsies for TM status and ultrasound body composition measurements of LM area (LMA), 12th rib fat thickness (FAT), and LM intramuscular fat percentage (IMF) for body composition were collected every 56 d. Data were analyzed as repeated measures using PROC MIXED with bull as experimental unit and fixed effects of TM, breed, time, and appropriate interactions. Results are presented as LSM ± pooled SE. All growth, body composition, and liver TM measurements were affected (P < 0.02) by time. There was no effect (P > 0.05) of TM source on body composition or liver TM concentrations. Inorganic had greater (P = 0.01) mean ADG (1.35 vs. 1.20 ± 0.04 kg/d) but lesser (P = 0.01) mean BCS (4.8 vs. 4.9 ± 0.03) than organic. Pooled across time, Brangus had greater (P < 0.05) LMA (64.95 vs. 60.43 ± 1.46 cm2), HH (119.3 vs. 114.6 ± 0.65 cm), BW (359 vs. 340 ± 6.0 kg), and BCS (4.9 vs. 4.8 ± 0.03) but lesser (P < 0.05) IMF (2.70 vs. 3.60 ± 0.14), than Angus, respectively. Liver Cu and Se were affected by breed × time (P < 0.05). On d 0, breeds were similar (P > 0.05) for Cu (109 ± 24.3 μg/g) and Se (0.94 ± 0.06 μg/g). Whereas, at d 168, liver Cu (297 vs. 148 ± 26.1 μg/g) and Se (1.76 vs. 1.42 ± 0.07 μg/g) were greater (P < 0.05) in Brangus than Angus, respectively. Prenatal and postnatal TM source had no effect on liver TM status of bulls. [ABSTRACT FROM AUTHOR]

A 2 × 2 factorial design evaluated breed (Angus vs. Brangus) and prenatal/postnatal TM source (inorganic vs. organic) effects on bull sexual development. Bulls (241 ± 2 d, 249 ± 4 kg, n = 32, 8 per TM × breed) born to dams supplemented with Co, Cu, Mn, Se, and Zn as inorganic (Se selenite or salt sulfates) or organic (Se-yeast and proteinates) TM sources starting 82 ± 2 d pre-calving. At weaning, calves were assigned to 1 of 4 pens based on treatment, receiving the same TM supplement as their dams. All bulls were fed a high energy/ protein diet to gain approximately 1.14 kg/d with TM source delivered in a pellet based on manufacturers recommendations. Bi-weekly semen collection, scrotal circumference (SC) and BCS (scale 1 to 9) were recorded. Semen collection was initiated when SC was 26 cm. Puberty was defined as an ejaculate with sperm concentrations ≥ 50 × 106 cells/mL, with ≥ 10% motility. Sexual maturity was defined as two consecutive semen collections with SC ≥ 30 cm, sperm motility ≥ 30%, and normal sperm ≥ 70%. Data were analyzed with PROC MIXED with fixed effects of TM source, breed, and interaction for puberty and sexual maturity data. Results are presented as LSM ± SE. At puberty (n = 27), no effect (P > 0.05) of TM source, breed, or interaction for age (337 ± 7 d), BCS (4.8 ± 0.1), motile (33.0 ± 2.3%) or normal sperm (48.7 ± 4.5%) occurred. However, Brangus (172.4 ± 28.2 × 106 cells/mL) had greater (P = 0.05) sperm concentration than Angus (96.9 ± 23.3 × 106 cells/mL). A TM source × breed effect (P = 0.01) occurred for SC at puberty; Brangus inorganic (33.5 ± 0.9 cm) were greater (P < 0.05) than Brangus organic (30.3 ± 0.9 cm), while Angus inorganic (29.9 ± 0.7 cm) and organic (31.2 ± 0.7 cm) were similar (P > 0.05). At sexual maturity (n = 15), organic (351 ± 18 d) were numerically (P = 0.14) younger than inorganic (391 ± 16 d), while BCS (5.2 ± 0.1), SC (33.6 ± 0.7 cm), sperm concentration (156.5 ± 24.9 × 106 cells/mL), motile (47.3 ± 2.7%), and normal (76.4 ± 1.0%) did not differ (P > 0.05) by TM source, breed, or interaction. Pubertal traits were affected by breed but not TM source. However, TM source may decrease bull age at sexual maturity. [ABSTRACT FROM AUTHOR]

Winter and spring born beef calves (n=568; 177.7 kg ± 34.0) at three different locations in Florida (MAR, BRU, SFE) were used to determine the effectiveness of deworming prior to weaning in order to achieve heavier weaning weights. At least three breed types were available at each location and included Angus, Brangus, Brahman, and Romosinuano. Calves were individually weighed unshrunk on d 0 and randomly assigned to treatment groups. Unshrunk BW were also obtained at a mid-point and on d 90. Treatments included no deworming (CON) or deworming (DW; doramectin injectable, 1 mL / 50 kg BW). Bahiagrass (Paspalum notatum) pastures were grazed at all locations; within a location, cow-calf pairs from both treatment groups were managed identically and forage allowances were monitored to minimize any potential bias due to variations in pasture conditions. Data were analyzed using Proc GLM and Proc Mixed of SAS. In the GLM procedure, data were blocked by location and treatment differences were tested using location x treatment as the error term. In Mixed, an analysis of covariance was performed with d 0 BW as the covariate; the Mixed model contained treatment as a fixed effect and the random effect was location. Within location, total gain and ADG was not different. Across all locations, DW calves gained more total weight (P < 0.001; 59.5 kg vs 55.0 kg) and ADG was greater (P < 0.001; 0.66 kg/d vs 0.62 kg/d) compared with CON. Although weight gain was greater (P = 0.002; 38.3 kg vs 35.5 kg) for DW calves during period 1, the difference was not as great during period 2 (P = 0.06; 21.1 kg vs 19.9 kg). Deworming costs averaged $1.57/hd. DW calves returned $1.94/hd more net revenue than CON calves when considering only the deworming product costs. Under these experimental conditions, our data indicated both an animal performance advantage and a positive ROI from deworming early. This management practice may improve the value of calves beyond that of simply more weight at weaning. Providing a healthier calf to the stocker or feedlot with process verification of this practice may garner additional premiums. [ABSTRACT FROM AUTHOR]

Background: The genes encoding β-actin and GAPDH are two of the most commonly used reference genes for normalization in in vitro blood-brain barrier studies. Studies have, however, shown that these reference genes might not always be the best choice. The aim of the present study was to evaluate 10 reference genes for use in mRNA profiling studies in primary cultures of brain endothelial cells of bovine origin., Methods: Gene evaluations were performed by qPCR in mono-culture and in co-cultures with astrocytes. The expression of reference genes was furthermore investigated during culture. Qbase+ software was used to analyze the stability of the tested genes and for determinations of the optimal number of reference genes., Results: The stability of the reference genes varied between the culture configurations, but for all culture configurations we found that the optimal number of reference genes were two. PMM-1, RPL13A and β-actin were the most stable genes in mono-cultures, non-contact co-culture and contact co-culture respectively. For studies comparing gene expression between different culture configurations the optimal number of reference genes was three and RPL13A was found to be most stable. During cell culture a number of four reference genes were found to be optimal and YWHAZ was found to be the most stable gene. β-actin and GAPDH were found to be the least stable genes during culture., Conclusion: Overall we found that the validation of reference genes was important in order to normalize target gene expression correctly, and suggest sets of reference genes to be used under different experimental conditions, in order to quantify mRNA transcript levels in blood-brain barrier cell models correctly., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Insulin and its receptor are known to be present and functional in the brain. Insulin cerebrospinal fluid concentrations have been shown to correlate with plasma levels of insulin in a nonlinear fashion, indicative of a saturable transport pathway from the blood to the brain interstitial fluid. The aim of the present study was to investigate whether insulin was transported across brain endothelial cells in vitro via an insulin receptor-dependent pathway. The study showed that the insulin receptor was expressed at both the mRNA and protein levels in bovine brain endothelial cells. Luminally applied radiolabeled insulin showed insulin receptor-mediated binding to the endothelial cells. This caused a dose-dependent increase in Akt-phosphorylation, which was inhibited by coapplication of an insulin receptor inhibitor, s961, demonstrating activation of insulin receptor signaling pathways. Transport of insulin across the blood-brain barrier in vitro was low and comparable to that of a similarly sized paracellular marker. Furthermore, insulin transport was not inhibited by coapplication of an excess of unlabeled insulin or an insulin receptor inhibitor. The insulin transport and uptake studies were repeated in mouse brain endothelial cells demonstrating similar results. Although it cannot be ruled out that culture-induced changes in the cell model could have impaired a potential insulin transport mechanism, these in vitro data indicate that peripheral insulin must reach the brain parenchyma through alternative pathways rather than crossing the blood-brain barrier via receptor mediated transcytosis.

Nearly 20 years have passed since the US Food and Drug Administration (FDA) approved the first companion diagnostic and today this type of assay governs the use of 21 different anticancer drugs. The regulators deem these assays essential for the safe and effective use of a corresponding therapeutic product. The companion diagnostic assays are important both during the drug development process as well as essential treatment decision tools after the approval of the drugs., (© 2017 American Society for Clinical Pharmacology and Therapeutics.)

Over the last couple of decades, molecular diagnostics have played an increasing role in drug development. Especially within oncology, more and more drugs are being developed together with a predictive biomarker assay using the drug-diagnostic codevelopment model. Not only do these assays support the development process but also the use of the drugs after regulatory approval as an important treatment decision tool. When these predictive biomarker assays are linked to a specific drug, they are called companion diagnostics. Furthermore, these assays are also considered an important element in the realization of precision medicine. Today, 21 different drugs have obtained US FDA approval together with a companion diagnostic assay, and the requirement for testing is part of their regulatory labeling. More than half of these drugs are for treatment of non-small cell lung cancer (NSCLC). With the approval of the different programmed cell death 1 (PD-1)/programmed cell death ligand 1 (PD-L1) immune checkpoint inhibitors, for the treatment of advanced stage NSCLC, a new class of predictive biomarker assays-complementary diagnostics-has emerged. Until now, 3 immune checkpoint inhibitors have obtained regulatory approval for treatment of NSCLC, and they all have a biomarker assay linked to their use. However, only for pembrolizumab, the PD-L1 immunohistochemical (IHC) 22C3 pharmDx assay has status as a companion diagnostic. For nivolumab and atezolizumab, the assays PD-L1 IHC 22C3 pharmDx and Ventana PD-L1 (SP142) have status as complementary diagnostics, which means that there are no requirements for testing included in the labeling for these drugs. Here, the authors discuss the clinical performance of the different IHC PD-L1 expression assays including the selection of the clinical cutoff values.

The variability of pharmacotherapy can be of a significant magnitude, and the main reason for this is often diseases heterogeneity. Patients who have similar diagnoses very often respond differently to the same pharmacological intervention, with great variability in both efficacy and safety outcome. Despite having discussed personalized medicine for more than a decade, we still see that most drug prescriptions for severe chronic diseases are largely based on 'trial and error' and not on solid biomarker data. However, with the advance of molecular diagnostics and a subsequent increased understanding of disease mechanisms, things are slowly changing. Within the last few years, we have seen an increasing number of predictive biomarker assays being developed to guide the use of targeted cancer drugs. This type of assay is called companion diagnostics and is developed in parallel to the drug using the drug-diagnostic co-development model. The development of companion diagnostics is a relatively new discipline and in this review, different aspects will be discussed including clinical and regulatory issues. Furthermore, examples of drugs, such as the ALK and PD-1/PD-L1 inhibitors, that have been approved recently together with a companion or complimentary diagnostic will be given., Competing Interests: Conflicts of Interest: Jan Trøst Jørgensen has worked as a consultant for Dako, Agilent Technologies and Euro Diagnostica, and has given lectures at meetings sponsored by AstraZeneca, Merck Sharp & Dohme, and Roche. Maria Hersom has nothing to declare. All the figures included in the manuscript are original and have been developed by Dr. Hersom.

Receptor-mediated transcytosis of the transferrin receptor has been suggested as a potential transport system to deliver therapeutic molecules into the brain. Recent studies have however shown that therapeutic antibodies, which have been reported to cross the brain endothelium, reach greater brain exposure when the affinity of the antibodies to the transferrin receptor is lowered. The lower affinity of the antibodies to the transferrin receptor facilitates the dissociation from the receptor within the endosomal compartments, which may indicate that the receptor itself does not necessarily move across the endothelial cells by transcytosis. The aim of the present study was to investigate transferrin receptor expression and role in transendothelial transferrin transport in cultured bovine brain endothelial cell monolayers. Transferrin receptor mRNA and protein levels were investigated in endothelial mono-cultures and co-cultures with astrocytes, as well as in freshly isolated brain capillaries using qPCR, immunocytochemistry and Western blotting. Transendothelial transport and luminal association of holo-transferrin was investigated using [ 125 I]holo-transferrin or [ 59 Fe]-transferrin. Transferrin receptor mRNA expression in all cell culture configurations was lower than in freshly isolated capillaries, but the expression slightly increased during six days of culture. The mRNA expression levels were similar in mono-cultures and co-cultures. Immunostaining demonstrated comparable transferrin receptor localization patterns in mono-cultures and co-cultures. The endothelial cells demonstrated an up-regulation of transferrin receptor mRNA after treatment with the iron chelator deferoxamine. The association of [ 125 I]holo-transferrin and [ 59 Fe]-transferrin to the endothelial cells was inhibited by an excess of unlabeled holo-transferrin, indicating receptor mediated association. However, over time the cell associated [ 59 Fe]-label exceeded that of [ 125 I]holo-transferrin, which could indicate release of iron in the endothelial cells and receptor recycling. Luminal-to-abluminal transport of [ 125 I]holo-transferrin across endothelial cell monolayers was low and not inhibited by unlabeled holo-transferrin. This indicated that transendothelial transferrin transport was independent of transferrin receptor-mediated transcytosis., (Copyright © 2016 Elsevier Inc. All rights reserved.)