H3Africa partnerships to empower clinical research sites to generate high-quality biological samples

Background The Institute of Human Virology Nigeria (IHVN) – Human Heredity and Health in Africa (H3Africa) Biorepository (I-HAB) seeks to provide high-quality biospecimens for research. This depends on the ability of clinical research sites (CRS) – who provide biospecimens – to operate according to well-established industry standards. Yet, standards are often neglected at CRSs located in Africa. Here, I-HAB reports on its four-pronged approach to empower CRSs to prepare high-quality biospecimens for research. Objectives I-HAB sought (1) to assess a four-pronged approach to improve biobanking practices and sample quality among CRSs, and (2) to build human capacity. Methods I-HAB partnered with two H3Africa principal investigators located in Nigeria and Ghana from August 2013 through to May 2017 to debut its four-pronged approach (needs assessment, training and mentorship, pilot, and continuous quality improvement) to empower CRSs to attain high-quality biospecimens. Results Close collaborations were instrumental in establishing mutually beneficial and lasting relationships. Improvements during the 12 months of engagement with CRSs involved personnel, procedural, and supply upgrades. In total, 51 staff were trained in over 20 topics. During the pilot, CRSs extracted 50 DNA biospecimens from whole blood and performed quality control. The CRSs shipped extracted DNA to I-HAB and I-HAB that comparatively analysed the DNA. Remediation was achieved via recommendations, training, and mentorship. Preanalytical, analytical and post-analytical processes, standard operating procedures, and workflows were systematically developed. Conclusion Partnerships between I-HAB and H3Africa CRSs enabled research sites to produce high-quality biospecimens through needs assessment, training and mentorship, pilot, and continuous monitoring and improvement.

http://www.ajlmonline.org Open Access an aliquot of all DNA. The bioinformatics core stored data. Beiswanger et al. 10 described the process for accessing biospecimens.
The NIH funded the Institute of Human Virology Nigeria H3Africa Biorepository (I-HAB). I-HAB's primary pursuit in Phase I was to achieve international standards according to the International Society for Biological and Environmental Repositories (ISBER) best practices. 11 For example, ISBER provides requirements for biospecimen collection, processing, quality control (QC), storage and transport, infrastructure, quality management, safety, training, and a laboratory information management system (LIMS). 11 The LIMS documents and tracks biospecimen attributes from collection through to use, depletion, and destruction. I-HAB also referred to the International Organization for Standardization (ISO) 15189 standards for clinical laboratories (https://www. iso.org/standard/56115.html) and Strengthening Laboratory Management Toward Accreditation (SLMTA) (https://slmta. org) for requirements pertaining to quality management, as appropriate. Regarding specific protocols, I-HAB referred to manufacturer specifications and journals.
Considering the potential imbalance between standardised preanalytical processes according to ISBER, ISO, and SLMTA and the lack of such standards in laboratories in Africa, I-HAB sought to evaluate assigned clinical research sites (CRSs) and implement improvement strategies. I-HAB established a training and mentorship programme to bridge gaps and to ensure researchers obtain biospecimens of the highest quality. This article describes I-HAB's experience in engaging two H3Africa projects from August 2013 through May 2017 in preparation for Phase II. 10

Methods
The NIH assigned I-HAB six research projects including Project A -a CRS located in Tanzania (not included in this study), Nigeria, and Ghana, with the project's central laboratory hub (clinical research sites sends biospecimens to laboratory hubs for processing, testing or storage) in Ghana -and Project B, with a CRS and laboratory hub in Nigeria. I-HAB employed a four-pronged approach of needs assessment, training and mentorship, piloting, and continuous quality improvement to help sites to collect, process, store, and transport high-quality biospecimens for future research.
A working group of NIH experts and representatives from each H3Africa biorepository developed guidelines and protocols to guide the H3Africa consortium on biorepository processes for harmonisation and consistency. The documents included biospecimens deposit and access requirements, and specific procedures for biospecimen collection, processing, transport, and shipping (www.h3africa.org). For example, the group established minimal criteria for DNA purity and concentration to ensure quality and consistency. The guidelines required submitters to maintain: • Ethical standards: ethical approval, informed consent, and Material Transfer Agreements (MTA).
• Legal requirements: shipping regulations and import and export permits. • H3Africa requirements: DNA quality, minimal data set, shipping checklist, and manifest and query forms. The 'minimal data set' included a biospecimen identification code, de-identified participant identification code, study name, specimen type, date of collection, gender, age at collection, and storage box position. The working group standardised the format of each element to ensure consistency and decrease errors.
The working group based the contents of the standard operating procedures (SOPs) on International Air and Transport Association regulations, ISBER best practices, manufacturer's specifications, and well-established practices as specified. ISO 15189 standards were useful for creating SOPs and documents regarding quality management and for defining the structural elements required for all SOPs. I-HAB recommended that sites customise SOPs to their laboratories.

Ethical Considerations
This article followed all ethical standards for research without direct contact with human or animal subjects.

Needs assessment
The I-HAB and research teams met to discuss project descriptions, processes, and procedures that potentially impact a biospecimen's quality. I-HAB reviewed study protocols and other documents in consideration of responsibilities and procedures impacting a biospecimen's integrity. I-HAB assessed facilities, equipment, personnel, documents and records, specimen management, organisation, purchasing, inventory, and quality assurance measures in line with underlying principles of ISBER best practices and ISO 15189 to ensure the sites could meet protocol requirements according to industry standards. Assessments included a combination of discussion, SOP review, site observation, and competency assessments for specifically assigned tasks. I-HAB offered document development, training and mentorship, biospecimen storage, distribution, and QC to resolve problems identified by assessments.

Continuous quality improvement
I-HAB engaged sites in continuous quality improvement. Continuous improvement is a cyclical process to improve quality by identifying problems and resolving them. I-HAB reviewed biospecimen QC, and shipment packaging, temperature, and documentation. I-HAB provided additional training or pilot exercises where appropriate to address nonconformities. The teams implemented corrective and preventive measures before initiating Phase II shipments to I-HAB. I-HAB determined improvement by the elimination of previously identified problems, improvements in DNA quality, and overall improvements in efficiency.
Following pilot deposition at I-HAB, Project A and Project B continued to identify opportunities for improvement. Project hubs monitored temperatures in shipments between I-HAB and project hubs. I-HAB monitored biospecimens and corresponding data for all subsequent deposits and communicated discrepancies to the sender within one week of discovery. Similarly, hubs continued to QC 100% of the DNA deposited. To ensure accuracy, I-HAB re-tested 10% of the DNA received. If more than 10% of the re-tested DNA was beyond acceptable range or inconsistent with I-HAB's results, then I-HAB tested the remaining 90% (total 100%). In such instances, I-HAB tested 100% of subsequent batch shipments from the project, until the project completed two consecutive shipments for which the QC for at least 90% of the DNA tested was within acceptable range and consistent with I-HAB's results. I-HAB continued to meet with the projects' staff to discuss future activities, challenges, and strategies of improvement.

Results
Engagement activities occurred over the three years of Phase I (2013-2016), in preparation for Phase II. I-HAB observed improvements in staff proficiency, procedural efficiency, and supply upgrades. I-HAB trained 51 persons in over 20 topics. I-HAB further determined improvements in supplies by the elimination of previously identified issues that resulted in poor sample quality. Preanalytical, analytical, and post-analytical processes, documents, and workflows were systematically created and modified based on needs assessment, training, mentorship, pilot exercises, and continuous quality improvement.

Project A
The study's investigators established a system for Phase II. The CRS in Nigeria collected, processed, temporarily stored, and shipped biospecimens to the project's hub in Ghana. An independent, commercial contractor extracted and QC tested DNA. The hub stored DNA and shipped the aliquots required for deposition to I-HAB. I-HAB performed needs assessment, training, and the pilot of the entire study process with the site in Nigeria, then piloted DNA shipment from Ghana to I-HAB. The process of engagement with Project A's CRS in Nigeria is summarised in Figure 1.

Needs assessment
In March 2014, I-HAB's manager, supervisor and QC officer assessed the CRS in Nigeria to compare infrastructural, commodity, and staffing resources to the laboratory protocol, H3Africa requirements, and best practices according to ISBER. The staff included an entry-level laboratory technician and data clerk. I-HAB evaluated the laboratory, staff, equipment, supplies, procedures, and workflows. The I-HAB manager communicated assessment outcomes and recommendations to the site principal investigator, study coordinator, laboratory technician, and data clerk as appropriate. I-HAB and Project A agreed that training, mentorship, and document development were required for Phase II preparation.

Training and mentorship
I-HAB provided training and mentorship for Project A's clinical site laboratory technician and data clerk: • The laboratory technician attended a five-day workshop at I-HAB, September 2013 (Table 1). • I-HAB provided customised five-day training for the laboratory technician, incorporating her responsibilities per the study protocol and H3Africa specifications, January 2014 (Table 1). • The I-HAB manager, supervisor, and biospecimens QC officer mentored the laboratory technician and data clerk for one day at their facility during a dry run on February 2014. ▪ The dry run simulated activities from participant arrival through to mock biospecimen collection, processing, temporary storage, and transport, including documentation. During the two-day dry run, improvements maximised client and staff safety, specimen integrity, and efficiency. I-HAB rearranged the biospecimen collection and processing area to reflect the natural flow of activities. Samples were inadequately separated during processing, and storage vials popped once they were frozen. Thus, I-HAB made recommendations that improved biospecimen integrity: (1) upgrade the centrifuge to enable staff to process biospecimens with adequate centrifugal force; (2) separate and freeze plasma, red cells, and buffy coat immediately, rather than freeze whole blood in Ethylenediaminetetraacetic acid vacutainers without processing them, and (3) (Table 1). Trainees were five laboratorians, one coordinator, two nurses, and four data clerks. Nine (90%) trainees who took pre-tests and post-tests increased in performance (Figure 2). The average pre-test scores were 62% (range 30% to 80%) while average post-test scores were 84.4% (range 63% to 100%). As expected, laboratorians performed better than non-laboratorians on average.

Project A, hub site (Ghana)
During Project A's hub's pilot in September 2015, Project A shipped 20 unique DNA biospecimens (extraction and QC at a commercial laboratory) to I-HAB by DHL on dry ice in four days. Ghana transferred data via a template comma-separated values file. The DNA was still frozen upon receipt, but temperature monitors were not included in the shipment. Also, there were inconsistencies between site versus biorepository results. Only 5% of the DNA was within the acceptable purity range per Ghana's result versus 25% per I-HAB ( Table 2). The average purity reported by both was below the acceptable standard. Furthermore, DNA concentration averaging 2.0 ng/μl (0.1-7.1) was lower than the recommended for deposition by H3Africa consortium. NanoDrop 2000 and 8000 series have lower detection limits of 2 ng/μl and 2.5 ng/μl; thus, measurements below these limits are unreliable. The DNA was not evaluated by agarose gel electrophoresis due to poor DNA integrity. These samples were tagged as low quality before storage and results were communicated and discussed with Project A. The major benefits of the pilot were knowledge of Project A's contractor's poor DNA quality and noncompliance with the shipping procedures.

Continuous quality improvement
Through remediation, problems in DNA concentration and purity, and temperature monitoring improved as observed from Project A's hub's subsequent shipment to I-HAB.

Project B (Nigeria) Needs assessment
Project B staff and I-HAB teams met three times: May, July and August 2014. The meetings facilitated an understanding of each other's goals, requirements, expectations, and services. Consequently, Project B requested that I-HAB introduce biospecimens management during its staff orientation (Figure 3). The I-HAB manager and supervisor reviewed Project B's study protocol to determine appropriate training topics.

Training and mentorship
The I-HAB supervisor attended Project B's five-day studywide orientation in July 2014, which included 35 research staff. The supervisor facilitated a biospecimen management workshop for one day, which spanned 14 topics (Table 3). She also reviewed supplies required for biospecimen collection, processing, and storage, and made recommendations, such as for laboratory supplies. For example, I-HAB suggested supplies for shipment of biological samples, Freezerworks basic LIMS software for sample management, and software for remote temperature monitoring and alerts, which remain in use by Project B. The I-HAB supervisor trained the project analyst in Freezerworks customisation and operations over five days, in late July 2014. Consequently, the analyst successfully customised the software, and adopted Freezerworks as the LIMS for biospecimen management.

Pilot
In January 2015 I-HAB drafted a protocol outlining pilot procedures. The teams attained ethical, legal, and regulatory requirements for the pilot. The MTA took one month to execute.

Score (%)
Pre-test % Post-test % Lab, laboratory. In June 2015 Project B extracted, QC tested, and shipped biospecimens to I-HAB by road via TRANEX (Lagos, Nigeria) in one day. There were two shipments: one on dry ice (DNA-5, Plasma-4, and serum-5) and one at a controlled ambient temperature (DNA-5). I-HAB conducted DNA QC as specified above. Outcomes and recommendations were communicated.

I-HAB Project A I-HAB Project A I-HAB Project A I-HAB Project A I-HAB Project B I-HAB Project B I-HAB
Project B I-HAB Project B The pilot identified areas that required improvement before Phase II. The hub forgot to send the manifest and the query form prior to the shipment. Both shipments contained insufficient refrigerant and were warm. The qualitative temperature indicators were not activated prior to shipment and the quantitative temperature loggers were misplaced. The logger for controlled ambient temperature was placed in the frozen shipment, while the logger for frozen shipments was placed in the controlled ambient temperature shipment. Nevertheless, 100% of the DNA had acceptable purity and concentration ( Table 2). The benefit of the pilot was to learn of issues with biospecimen shipping procedures.

Continuous quality improvement
I-HAB provided remedial training for the laboratory manager and study coordinator in July 2015 (

Discussion
Researchers inexperienced with processes that influence biospecimen integrity may attain high-quality biospecimens for research by partnering with a biorepository and establishing a system of needs assessment, training and mentorship, pilot, and continuous quality improvement that is based on best practices. It is beneficial to start with general meetings and needs assessments to understand the intended goals and procedures and whether resources and capabilities are well aligned. Even if a scientist or contractor claims to be to be proficient, they should be subjected to a standardised assessment to ensure competency. Had Project A subjected its contractor to assessment, it would have learned of the contractor's poor performance prior to study activation. The contractor had not resolved issues with purity that were indicative of protein contamination (purity ratio below 1.7) as of the first Phase I shipment. Likewise, Project B's new hire experienced issues with DNA quality that could be circumvented via an assessment. Both examples demonstrate that assumptions of competency risk financial and biological resources. They also demonstrate the imperativeness of training and monitoring.
The key to effective training is to establish a foundation of harmonised guidelines, procedures, and minimal requirements that align with project goals and best practices. The biorepository should utilise standard and customised training that encompasses theory, practical exercises, and post-tests. Training achieved staff competency and eliminated issues revealed by needs assessments, pilot exercises, and continuous monitoring. Both projects improved in performance following training; however, biological shipping may require refresher training or additional practical exercises. Both projects also gave public testimonials at scientific meetings that prompted interest and trust of other researchers.
The pilot process allows biorepositories to identify challenges in the cascade of processes leading to biospecimen deposit that may only be revealed by a real-time trial that involves all direct and indirect processes. For example, the pilots identified and eliminated errors in shipping that can only be unmasked by full trial. Due to the peculiarities of packaging and International Air Transport Association regulations associated with international biological shipments, on-site mentorship is useful to reinforce procedures learned during training, according to the trainees' true environment and resources. Consequently, I-HAB incorporated site visits to assist sites to execute their first shipment to I-HAB.
Our outcomes mirrored the successes and challenges observed in other laboratory development programmes in sub-Saharan Africa that incorporate assessments, training, and mentorship. Generally, training and mentorship improved laboratory performance and empowered a new cadre of professionals. 14,15 Problems in biospecimen acceptability, 3 documents and records, information management, and supplies were common; however, training and mentorship facilitated improvements. 16,17 Literature suggests that relapses may occur after training, particularly in documentation. 14 Thus, I-HAB's method of continuous improvement shall be critical to maintain improvements.

Conclusion
Biobanking in Africa is expected to increase and improve with greater knowledge and resources. 6 However, CRSs with abilities to meet best practices to collect, process, and store biospecimens of good integrity and quality are scarce. Partnerships between competent regional biorepositories and research investigators may build human capacity and improve biobanking practices and biospecimen quality by adopting an approach of needs assessment, training and mentorship, pilot, and continuous quality improvement. The improvements in quality may improve performance, increase staff morale, and contribute to sustainability of the biobank, research entity, and biobanking industry.

Lessons learned
• Clinical research sites and biorepositories may partner to empower clinical research sites to generate high-quality biospecimens via needs assessment, training, pilot, and continuous quality improvement.
• It is critical to standardise guidelines, procedures, and minimal criteria early on to inform training and activities.
• Standardised guidelines, procedures, and minimal criteria should be rooted in wellestablished requirements such as ISBER best practices, ISO 15189, and SLMTA.