Allografts and Storage: What are the Temperature Requirements?

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Allografts are produced in different formats. The temperature and method of storage is dependent on the type of tissue, its method of preservation, and its intended use. Methods of cold storage for tissue include refrigerated, frozen, or frozen-cryopreserved techniques, while simple ambient room temperature is appropriate for lyophilized, dehydrated, or desiccated tissues. 1

Standards have been set for cold storage of tissues which identify acceptable temperature ranges and their impact on shelf life. Tissues that can be kept refrigerated (above freezing, 0°C to 10°C) most commonly include placental tissues, cellular tissues, and osteoarticular or osteochondral grafts. The shelf life will vary based on the tissue and processing method. Placental tissue storage temperatures are currently established by the processing tissue bank. 1

Frozen and cryopreserved allografts must be stored at ultra-low temperatures to maintain their viability. Ultra-low freezer temperatures of -20°C to -39°C generally allow a shelf life of 6 months, whereas -40°C to -86°C allow longer terms up to 5 years. For example, tendons, skin, and osteoarticular grafts can be stored at -20°C to -39°C for 6 months or less, and at -40°C to -86°C (or colder) for up to 5 years. 1

Frozen and cryopreserved tissues are not the same. Cryopreserved tissues are specially preserved with a solution containing a cryoprotectant, such as glycerol or dimethylsulfoxide. 1 The purpose is to prevent cell death during freezing and thawing. Cryopreserved grafts are frozen slowly at a controlled rate and are used in procedures requiring viable cells such as chondrocytes.

Cardiac and vascular tissues are cryopreserved and can only be stored at -100°C or colder. 1 Reproductive tissues such as semen and ova must be stored in liquid nitrogen, in either the liquid or vapor phase. Cellular and birth tissues have no FDA-established standards, and as such the temperatures are established by the tissue bank. 1

Lyophilized tissue, commonly referred to as “freeze dried tissue,” is tissue that has been dehydrated for storage by converting the water to gas under a vacuum that extracts moisture. 1 Dehydrated and dessicated tissues are created by removing the water from the tissue, accomplished through chemical means such as alcohol soaks, critical/supercritical drying, simple air drying, or drying in a desiccator. 1 Lyophilized, dehydrated, and dessicated tissues can be stored at ambient room temperature. 1

A temperature monitoring system is required for both refrigerated and frozen systems, which must be utilized to document temperatures and to alert staff with audible and/or visual alarms when temperatures stray outside of set acceptable limits. 1 Many modern freezers and refrigerators also have alarms for power outages, low battery, door opening, filter blockages, or system failures.

Standards of procedure require operational protocols to be in place for reviewing temperature logs, and documenting the review. When storage methods utilize liquid nitrogen cooling, monitoring of either temperature or liquid nitrogen levels is necessary, and documentation is required at regular intervals. 1

References:
1. American Association of Tissue Banks. (2016). Standards for Tissue Banking (14th ed.). J. C. Osborne, K. G. Norman, T. Maye, P. Malone, S. A. Brubaker, et al. (Eds.). Bethesda: Naval Medical Research and Development Command. Available from:
http://www.aatb.org/AATB-Standards-for-Tissue-Banking

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Good Tissue Bank Standards of Practice

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What helps make a tissue bank good?

Following Standards of Practice (SOPs) in tissue banking is vital to ensure that every step of human tissue procurement, preparation, and distribution is held to the highest standards. This is done to ensure the safety, quality, and traceability of tissues for surgical use.

The internal Standards of Practice for tissue banks include documents that cover every aspect of this process. These standards cover topics including monitoring tissue recovery sites, the audit of tissue recovery partners, necessary body cooling protocols, microbiological surveillance programs, providing service to tissue donor families, prevention of contamination and cross-contamination at recovery, and many more.

However, the most relevant guide to understanding what makes a “good tissue bank” are the factors that govern “Good Tissue Practice,” defined by the steps that the FDA defines in the manufacturing of human allografts for surgical use: recovery, donor screening and testing, packaging, labeling, distribution, processing, and storage.1 Detailed guidelines for these areas of “Good Tissue Practice” have been developed by the national organization which audits and accredits participating US tissue banks.2

Recovery of Allografts

Recovery is the process of obtaining from a donor either cells or tissues that are intended for human use via implantation, transplantation, infusion, or transfer.1 It is the responsibility of the donor recovery agency to have a program in place that ensures donor information is received, investigated, evaluated, documented, and shared with all agencies that receive tissues from the donor to ensure donor suitability. There are specific SOPs in place to ensure that all recovery personnel are well trained and competent.

There are very specific steps that are taken by the donor recovery operation to control contamination and cross-contamination during retrieval. These steps include adherence to appropriate donor eligibility guidelines such as specific body cooling parameters and time limits for retrieval; ensuring a suitable location for recovery site activities; using clean techniques appropriate to the specific cells/tissues being recovered; cleaning, disinfecting, and sterilization of equipment, supplies, and instruments; monitoring recovery activities for microorganism contamination, such as by culture results; and sharing of all records related to donor eligibility determinations.

Donor Screening and Testing

The main step to prevent potential infectious diseases in the donor tissue supply is primary prevention, which comes from donor screening and testing, first in the form of the detailed medical-social interview (Donor Risk Assessment) of the donor and exhaustive review of their medical records and sexual history. There are multiple levels of review of every record, including by an independent medical doctor specializing in cytopathology. All tissues are also required to be tested by a range of FDA-licensed, cleared, or approved donor screening tests, as applicable by tissue type, such as for HIV, HBV, HCV, syphilis, HTLV, CMV, etc.3 Testing laboratories must also be registered as tissue establishments by the FDA, and are subject to the same rigorous oversight as tissue banks and procurement organizations.

Processing and Storage

Processing is defined by the FDA as “any activity performed on an HCT/P, other than recovery, donor screening, donor testing, storage, labeling, packaging, or distribution, such as testing for microorganisms, preparation, sterilization, steps to inactivate or remove adventitious agents, preservation for storage, and removal from storage.”4 Generally, processing includes cutting, grinding, shaping, culturing, enzymatic digestion, and decellularization of recovered tissues to make them suitable for surgical use.3 Processing must be conducted in a suitable clean room environment with appropriate environmental controls and monitoring.

Processing may also include a sterilization step such as gamma or e-beam radiation. Both are validated, controlled methods of improving allograft tissue safety. Most tissue banks employ a sterilization step in the face of infectious disease risks. To learn more about sterilization through radiation, see this post: “Understanding Tissue Sterilization, Part 2: Radiation Round-up.”

Storage of allografts must be maintained at appropriate temperatures within each tissue’s tolerance limits. A temperature monitoring system must be in place to document storage temperatures and alert staff of deviations before straying outside of acceptable limits. Each unit of stored tissue should be packaged to facilitate sterile storage and prevent contamination or cross-contamination.

Packaging and Labeling

Packaging and shipping are not part of processing, and are therefore not required to be validated or verified in the same way as manufacturing. However, appropriate procedures must be in place to ensure the tissues are protected from contamination and cross-contamination, and that appropriate storage parameters are maintained during transport, particularly that temperature ranges are kept within acceptable limits.

Tissue labeling must make appropriate claims that are supported by verification and validation data to ensure that they are true and accurate. This could include information on sterilization procedures. Labeling also must include a method of internal donor identification, usually a lot-based identifying number on the label to allow for tracking and traceability of every tissue.

Distribution of Allografts

Strict regulations govern the transport and distribution of allografts, including an expiration date for their use. The expiration date is the maximum allowable storage period for the tissue, and expired tissues should not be transplanted. It is also critical to ensuring a safe allograft supply that the distribution of tissues be completely traceable from the original donor, to the retrieval agency, to the intermediary or processor, and on to any other tissue manufacturer or distributors to the ultimate end-user. The information that should travel with an allograft includes tissue ID numbers, tissue type, quantity, time of transport and delivery, recipient of delivery, and who transported and accepted tissues.

If you need help developing a tracking process for your surgical facility, there are resources available to guide you. In addition to the information above, a tissue tracking log should reflect the date of implantation and recipient of every graft. More information on tracking requirements can be found at the FDAs website.