By Alberta Colakovic, Brian Hewson, and Tal Murthy Thursday, April 1, 2024

2D Barcodes ensure that a multitude of samples can be tracked in a variety of storage conditions.

Sample identification is a vital process to all scientific laboratories. With the need to track and handle thousands, or even millions of individual samples, an effective method of high-throughput sample identification, retrieval and processing from storage is required. In order to facilitate this process, two dimensional (2D) barcoded tubes have been manufactured which feature unique 2D datamatrix codes on their base. This makes storing, retrieval and identification of various samples much quicker and easier for the user, ensuring that tubes can be readily tracked. As a result, the laboratory can effectively streamline its processes by eliminating mix ups and minimizing manual tracking operations, thus freeing up more time and resources for other experimental procedures. The use of 2D barcode tubes and some of the validation conditions are discussed here.

Introduction
Tubes that have been manufactured with a barcode are permanently tagged, providing the laboratory with a secure and reliable tracking method. 2D bar coding is more commonly used due to its ability to hold significantly more information than 1D barcodes. This not only provides an excellent method to monitor inventory, but also reduces the occurrence of sample misidentification. As a result, barcoded data provides the user with a level of assurance as to what is actually contained within each sample tube, thus eliminating the possibility of working with incorrect samples. Such errors may adversely affect the viability of some substances, and the addition of the wrong reagent to a reaction may result at best in inconsistent data and at worst in hazardous or detrimental effects. As such, using 2D barcoded tubes for laboratory sample management prevents mistakes, improves experimental reproducibility and safety, and helps to streamline laboratory operations.

The 2D datamatrix barcodes are permanently attached to the base of each storage tube, either via direct laser-etching or encapsulation, to ensure that viability is maintained for consistent readability. This provides both secure rack and tube traceability, regardless of the storage system in use. The information contained within the barcode can subsequently be translated by a complementary barcode reader and rapidly integrated with ease into sample storage archives such as spreadsheet-type databases or Laboratory Information Management Systems (LIMS).

Storage conditions
The ability to track and retrieve samples is imperative over long periods of time and can become seriously compromised if the barcode deteriorates. Different sample types are subjected to a variety of conditions and temperatures, depending on the best method for maintaining sample viability and integrity. Readability must therefore be maintained under all storage conditions. Typically, long-term storage of samples is carried out at -80°C, or increasingly, down to cryogenic temperatures in liquid nitrogen (approximately -196°C). Furthermore, some samples require ‘snap freezing’ in the vapor phase of liquid nitrogen prior to long term storage. Samples required for immediate experimentation or analyses are often stored at -4°C. As such the barcode needs to be resistant to a wide range of chemicals and abrasives as well as extremely low temperatures.

Ideally, samples should be aliquoted and stored in freezers as a library of viable samples to eliminate the chances of sample deterioration and potential contamination. Individual aliquots can then be thawed for immediate experimental use. However, this practice is not convenient for all laboratories—due to cost or space restrictions there may be situations where samples must be stored in larger aliquots. 2D barcodes, therefore, must be readable for samples that undergo freeze-thaw cycles.

2D barcoded tube testing
Three different Thermo Scientific tube types were subjected to different conditions to assess their ability to maintain readability of the 2D barcode: Matrix 1.4mL flat bottom, sterile; Nunc 1.0mL; Abgene 1.2mL twist-lock with caps. Each tube was tested under the following conditions: -80°C freeze; vapor phase liquid nitrogen; and boiling water. Although boiling water is not routinely used for the bar coded tubes, we wanted to use this extreme condition to ensure that the barcode is still functional under harsh conditions. Each tube rack used contained a column of eight tubes filled with the following liquids: 100% DMSO; distilled water; or empty.

The conditions are defined as follows: one freeze and thaw cycle at -80°C, the tubes were frozen in a -80°C freezer and thawed to room temperature; one freeze-thaw cycle in vapor phase liquid nitrogen, the tubes were frozen in liquid nitrogen and thawed to room temperature; and for one boiling cooling cycle, the tubes were boiled in 100°C water for 10 minutes and allowed to cool to room temperature. A tube thawing device was also used, which continuously blew ambient air over the bottom of the tubes to facilitate faster thawing. Once thawed, the racks of tubes were placed back into their respective storage conditions.

The 2D barcodes of filled tube racks were scanned twice using a complementary 2D barcode reader to establish optimal barcode readability in common laboratory conditions. The information was translated into numeric code and then stored as text files.

At -80°C, 950 μl and 75 μl aliquots typically undergo 10 freeze and thaw cycles. In vapor phase liquid nitrogen, although tubes can be sturdy, cells or tissues generally cannot withstand multiple freeze thaws; hence only five cycles were performed. In cases where repeated thawing of samples involving cells or tissues is required, it is advisable to store the samples in aliquots.

Each tube, therefore, underwent 10 freeze-thaw cycles at -80°C, five freeze-thaw cycles in liquid nitrogen and five boiling and cooling cycles. After freezing or boiling, the tube racks were allowed to thaw or cool to room temperature respectively, then read using the 2D barcode reader.

2D barcoded tube readability
This study clearly indicates that the datamatrix 2D barcoded tubes tested were able to withstand multiple freeze-thaw cycles, boiling and cooling cycles, as well as vapor phase liquid nitrogen, without showing any signs of degradation or any problems with bar code readability.

The results shown in Table 1 demonstrate that irrespective of content, 100% of the tubes from each tube rack were able to be interpreted into a numeric code after a total of 10 freeze-thaw cycles at -80°C, five freeze-thaw cycles in vapor phase liquid nitrogen, and five boiling and cooling cycles. Therefore, freezing and thawing, or boiling and cooling tubes numerous times, does not affect the barcode integrity of these three types of 2D barcoded tubes.

Conclusion
With an increasing need for storing and processing hundreds of thousands of samples under varying conditions, storage, and retrieval reliability is of paramount importance. Retaining barcode viability under common laboratory storage conditions is essential for the accurate traceability for large numbers of samples contained within any laboratory database or tracking system. As a part of a combined offering of sample storage solutions, Thermo Scientific 2D barcoded tubes maintain flawless readability after numerous cycles in a variety of storage conditions. The tubes exhibit datamatrix code durability, as well as code resilience and stability for optimal performance.

About the Authors
This study was performed at the Scientific Applications Laboratories of Thermo Fisher Scientific: The team Alberta Colakovic, Brian Hewson (brian.hewson@thermofisher.com), and Tal Murthy (tal.murthy@thermofisher.com) develop applications in different scientific fields on various products including the sample storage product lines.

This article was published in Bioscience Technology magazine: Vol. 34, No. 3, March, 2010, pp. 30-32.