The concept entered formal law through Directive 2010/63/EU, the European Union’s cornerstone legislation on the protection of animals used for scientific purposes, which calls on institutions to foster a climate of care. Guidance from the RSPCA, the Laboratory Animal Science Association and the EU’s own working documents on animal welfare bodies go further, defining a culture in which all staff, from principal investigators to animal technicians, are encouraged and enabled to take responsibility for animal welfare and ethical standards that go beyond the bare minimum the law requires. Since 2016, an International Culture of Care Network has connected institutions pursuing that ambition. What has been missing, the Utrecht team argues, is a clearly described, literature-based way to actually assess whether such a culture exists on the ground.

The study unfolded in two phases. The first was a scoping review conducted according to the PRISMA-ScR reporting guidelines, the standard framework for transparent scoping reviews. The researchers searched PubMed, Embase and Web of Science with a Boolean strategy combining terms for the Culture of Care and related concepts, such as caring culture, climate of care and organizational culture, with terms for the laboratory animal context, including animal research, in vivo and animal facility. The search, run on April 10, 2025, yielded 155 unique records after deduplication, which were screened independently and blinded by two reviewers. Twenty-one publications survived full-text screening, published between 2007 and 2024 with a median publication year of 2022, and drawn mostly from the United Kingdom, followed by the United States and Germany.

What the review revealed was striking. Most of the included articles described the Culture of Care at a purely conceptual level, and not a single one used a validated assessment instrument. Eleven articles included some form of assessment, six using surveys and five using qualitative methods such as interviews, but the approaches varied widely in scope and rigor. A quality assessment of the four survey-based studies, scored against simplified criteria drawn from the SURGE and CHERRIES guidelines, found that formal reliability reporting remained limited and bias control was generally modest. One study developed a 12-item scale evaluated with principal component analysis; others reported internal wellbeing surveys or questionnaires distributed through emails and mailing lists with little methodological detail. In short, the field had plenty of vision and very little instrumentation.

From the 21 publications, supplemented by policy documents from the EU and the International Culture of Care Network, the researchers distilled nine recurring thematic domains. Compliance with organizational and legal standards was the most universal, explicitly mentioned in 19 of the 21 articles, reflecting the view among many authors that meeting legal requirements is the starting point rather than the endpoint. Next came a proactive attitude that goes beyond compliance, mentioned in nine articles, followed by strong and caring leadership and sufficient resources and conditions to do a proper job, each named in 13 articles. Empathy and respect towards colleagues and animals, shared responsibility and accountability, and transparency and open communication each appeared in 15 to 17 articles, while a supportive work environment and team collaboration appeared in 16, and sharing knowledge and continuous reviewing in 15.

Several of these themes carry a distinctly human dimension that has gained prominence over the past decade. The Culture of Care builds on animal welfare science by emphasizing organizational culture and staff wellbeing, and the review noted that attention has shifted from animal care alone to the people who provide it. Compassion fatigue, the emotional exhaustion that arises from ongoing exposure to animal suffering, ethical tension and high workload, can erode vigilance, empathy and professional judgment, and thereby feed back into animal welfare. A related scoping review cited in the study found that the most common strategy suggested to mitigate work-related stress among laboratory animal personnel was acknowledgement and validation of their feelings. The new questionnaire reflects this dual focus, with items on feeling safe to share emotional reactions, knowing where to find support, and still having energy after a working day.

The second phase translated the nine themes into a measurable instrument. The researchers drafted 50 positively worded statements in the first person and present tense, each rated on a five-point Likert scale from strongly disagree to strongly agree, a design chosen to reduce cognitive load and encourage reflection on personal experience. Four questions were adopted directly from the NHS Culture of Care Barometer, a validated tool from human healthcare, using both the original English version and a published Dutch translation. Eleven staff members of the Animal Welfare Body Utrecht were invited to rate each item’s relevance and provide open-text feedback on clarity, redundancy and translation accuracy; eight responded, with mean relevance scores ranging from 3.8 to 4.8. After discussion with three experts, items with low relevance, overlap or vague wording were removed or merged, leaving a final 30-item questionnaire covering all nine themes.

The team then put the instrument through a test-retest study, asking participants to complete the same questionnaire twice, two weeks apart. The first round targeted 50 laboratory animal coordinators at Utrecht University and UMC Utrecht, of whom 15 completed the first round and 5 the second. An open call in October 2025 on MyDALAS, the platform of the Dutch Association for Laboratory Animal Science with roughly 700 active members, and on the International Culture of Care Network added 12 more test respondents, 11 of whom completed the retest. In total, 16 participants completed both timepoints. Internal consistency was very high at both timepoints, with Cronbach’s alpha of 0.917 at test and 0.898 at retest, well above the commonly cited acceptable range of 0.70 to 0.95, though values above 0.90 can hint at some redundancy among items.

Test-retest reliability, assessed with two-way absolute-agreement single-measure intraclass correlation coefficients, told a more nuanced story. The total-scale ICC was 0.73, with a 95 percent confidence interval of 0.38 to 0.90, which the authors characterize as moderately good, and the wide interval is expected given the small sample. At the level of individual items, ICCs ranged from poor, below 0.50, to good, between 0.75 and 0.90, with broad confidence intervals. The researchers explain this variability through classical test theory: single items carry more random error, while aggregated scores gain reliability as that error cancels out. They also identified ceiling effects, noting that one item was answered almost exclusively with agree or strongly agree at both timepoints, leaving too little variance between respondents to detect meaningful stability. Item-level results, they caution, should be interpreted with care.

The study’s limitations are candidly acknowledged. The two recruitment rounds differed in timing and incentives, with the second offering a randomly awarded 30 euro gift card, and participants recruited through a network already engaged with Culture of Care initiatives may have been more positively inclined toward the concept, introducing selection bias. Response rates could not be calculated because the number of people who saw the open calls is unknown, and no demographic data were collected to preserve anonymity in the small sample. Still, the authors argue, the work delivers what the field has lacked: a structured overview of how the Culture of Care is described, a nine-theme analytical framework, and a questionnaire with initial evidence of internal consistency and preliminary test-retest reliability. The next challenges are construct validity, factor structure, subgroup differences and performance in larger, more diverse samples, including whether the tool can detect change over time. If those steps succeed, institutions may finally be able to measure the invisible climate they have long been asked to cultivate.

Subject of Research: Development and testing of a Culture of Care questionnaire for laboratory animal institutions

Article Title: From concept to measurement: a scoping review and development of a Culture of Care questionnaire for laboratory animal institutions

Article References: Weber, O. M., van de Kolk, S. A. C., & Van Loo, P. L. P. (2026). From concept to measurement: a scoping review and development of a Culture of Care questionnaire for laboratory animal institutions. Frontiers in Ethology, 5, Article 1796584. https://doi.org/10.3389/fetho.2026.1796584

Image Credits: AI Generated

DOI: 10.3389/fetho.2026.1796584

Keywords: Culture of Care, laboratory animals, animal welfare, staff wellbeing, scoping review, questionnaire development, test-retest reliability, Cronbach's alpha, intraclass correlation, Directive 2010/63/EU, 3Rs, organizational culture

The conventional approach to freezing animal sperm, including rat sperm, typically involves a lactose-based solution supplemented with chicken egg yolk. Egg yolk earned its place in cryopreservation media decades ago because its lipoproteins help shield sperm membranes from the stresses of freezing and thawing. But it comes with baggage. As a biological product of variable composition, egg yolk introduces batch-to-batch inconsistency that makes it difficult to standardize freezing protocols between laboratories. More concerning is the risk of microbial contamination, since egg products can harbor bacteria and viruses that threaten both the samples being preserved and the animals that may eventually be produced from them. For colonies of laboratory rats maintained under strict pathogen-free conditions, that risk is far from trivial.

The second problematic ingredient is OEP, a detergent containing sodium lauryl sulfate that is commonly added to sperm freezing solutions to improve their performance. While OEP helps during the freezing process, it has a darker side when the thawed sperm are used for in vitro fertilization: the detergent has the potential to damage oocytes, the egg cells that sperm must fertilize. In an IVF setting, where the goal is to produce viable embryos from carefully collected eggs, introducing a substance that can harm those very eggs is an unwelcome compromise. These twin problems, microbial risk from egg yolk and oocyte toxicity from OEP, motivated the Kyoto University and RIKEN team to ask a fundamental question: could rat sperm be frozen effectively using only chemically defined components whose identity, purity, and concentration are fully known?

“We were particularly interested in whether rat sperm could be cryopreserved without egg yolk, which has been used for many years in conventional sperm preservation solutions,” says first author Kohtaro Morita of Kyoto University. The question matters because rats occupy a special place in biomedical research, serving as models for physiology, pharmacology, toxicology, and human disease. Yet the reproductive engineering technologies that have transformed mouse genetics, allowing researchers to archive and ship mouse strains as frozen sperm, have lagged behind for rats. A reliable, defined freezing solution would be a practical tool for maintaining rat colonies, exchanging genetic lines between institutions, and safeguarding valuable strains against accidental loss.

The team’s strategy was systematic and iterative. They began by freezing rat sperm in a simple lactose solution, stripped of both egg yolk and OEP, and then measured sperm motility after thawing. Motility, the ability of sperm to swim actively, serves as a key indicator of post-thaw viability and fertilizing potential. By varying the concentration of lactose across trials, the researchers identified the optimal level for supporting sperm survival through the freeze-thaw cycle. With that baseline established, they turned to additives, testing each candidate’s contribution to membrane protection and motility recovery.

The first meaningful improvement came from OptiPrep, a commercially available iodixanol-based density gradient medium that has previously shown effectiveness in protecting sperm during cryopreservation. When added to the lactose solution, OptiPrep improved sperm motility after thawing, suggesting it helps buffer the cells against the physical and osmotic stresses of ice formation. The researchers then layered on additional cryoprotectants in various combinations and concentrations, systematically searching for the formulation that yielded the highest motility in thawed rat sperm. This combinatorial screening approach allowed them to identify synergies between protective agents rather than relying on any single ingredient.

Two components emerged as particularly important. The combination of ethylene glycol, a small penetrating cryoprotectant widely used in embryo freezing, and sericin, a silk-derived protein with documented cell-protective properties, reduced damage to both the plasma membrane and the acrosomal membrane of the sperm. The acrosome is the enzyme-filled cap that sperm use to penetrate an egg during fertilization, so preserving its integrity is essential for maintaining fertilizing capacity. In addition, the researchers found that two nucleotides, ATP and dbcAMP, improved sperm motility after thawing, presumably by supporting the energy metabolism and signaling pathways that drive sperm movement. The final optimized solution thus consisted of OptiPrep, ethylene glycol, sericin, ATP, and dbcAMP in a lactose base, every component chemically defined and free of animal-derived biological material.

With the formulation in hand, the team put it to the ultimate test: could sperm frozen in this solution actually produce live animals? They cryopreserved rat sperm, thawed it, and used it for in vitro fertilization with fresh oocytes collected from female rats. The resulting two-cell embryos were then transferred into the oviducts of pseudopregnant female rats, where they could develop to term. To benchmark performance, the researchers ran the same IVF and embryo transfer procedure in parallel using freshly prepared, never-frozen sperm, allowing a direct comparison of fertilization efficiency and birth rates between the two groups.

The results were encouraging, if not perfect. Sperm cryopreserved in the new solution achieved a fertilization rate of approximately 70 percent, lower than the roughly 90 percent achieved with fresh sperm, but high enough to efficiently generate fertilized oocytes for embryo production. More striking was the outcome at the other end of the developmental pipeline: approximately 36 percent of the embryos fertilized with cryopreserved sperm resulted in live rat pup births, a figure that showed no significant difference compared with the fresh sperm group. In other words, once an embryo was successfully produced from the frozen-thawed sperm, its ability to develop into a healthy pup was essentially equivalent to that of an embryo derived from fresh sperm. The freezing process, in the new defined solution, did not appear to compromise developmental potential.

“Although reproductive engineering technology for rats has lagged behind that for mice, we expect that this new cryopreservation solution will facilitate the international transport of rat sperm and reduce the risk of microbial contamination when producing offspring,” says Morita. The implications extend across the laboratory animal science community. Shipping frozen sperm rather than live animals is cheaper, safer, and more humane, and it allows institutions to reconstitute strains locally from a small vial rather than transporting breeding pairs across borders. A defined, yolk-free medium also removes a persistent source of variability that has complicated comparisons between laboratories, since each batch of egg yolk carries its own biological idiosyncrasies.

The authors are candid that further improvements are still necessary. The fertilization rate achieved with the new solution, at around 70 percent, remains below the 80 to 95 percent range reported for fresh sperm or for conventional cryopreservation methods that include egg yolk. Closing that gap will likely require refining the formulation further, perhaps by tuning cryoprotectant concentrations or identifying additional membrane-stabilizing agents. Even so, the study, published on 16 September 2026 in Biology of Reproduction under the title “Development of a chemically defined cryopreservation solution for rat sperm and production of offspring by in vitro fertilization,” demonstrates that the long-standing compromises of conventional sperm freezing media can be overcome. By showing that rat sperm can be frozen without egg yolk or detergent, thawed, and used to generate healthy offspring through IVF, the Kyoto University and RIKEN team has opened a path toward cleaner, more standardized gamete banking, not only for rats but potentially for other species where the same old recipe has persisted for decades.

Subject of Research: Development of a chemically defined cryopreservation solution for rat sperm enabling in vitro fertilization and offspring production without egg yolk or detergent

Article Title: Rat sperm freezing without the egg yolk

Article References: Rat sperm freezing without the egg yolk. (n.d.). Original publication

Image Credits: AI Generated

DOI: Not provided

Keywords: cryopreservation, rat sperm, in vitro fertilization, egg yolk-free, sperm motility, ethylene glycol, sericin, OptiPrep, Kyoto University, RIKEN, reproductive technology, laboratory animals