Optimal Whole Blood Storage Conditions for Measuring BCR-ABL1 Accurately for Xpert^® BCR-ABL Ultra

Reliably quantifying BCR-ABL1 and ABL1 transcripts in whole blood for patients on TKI therapy is essential, but it can be challenging because intracellular RNA may rapidly degrade during storage. The purpose of the studies is to evaluate the effect of blood storage conditions with increased time interval on BCR-ABL1 and ABL1 gene expression, and demonstrate the validity of results in %BCR-ABL/ABL (IS) generated from Xpert^® BCR-ABL Ultra.

The whole blood stability was evaluated in contrived and CML patient samples. Contrived samples were prepared by spiking CML⁺ cells into normal EDTA or PAXgene blood at ~0.1% (IS). In EDTA, it was stable for 48 hours at room temperature (RT) and up to 3 days at 2-8°C. In PAXgene, it was stable for 5 days at RT or 2-8°C, and up to 25 months at -80°C. The CML EDTA specimens evaluated at ~10%, ~1%, ~0.1% and 0.01% (IS) were stable at RT for 48 hours and up to 4 days at 2-8°C.

To evaluate long term storage for whole blood, contrived samples at ~0.1% (IS) and CML EDTA specimens at various % IS were spotted onto a Dried Blood Spot (DBS) paper template. The DBS stability at RT was demonstrated for more than 40 days from the CML clinical specimens and up to 12 months on the contrived samples. In addition, the long term stability was evaluated on frozen lysates prepared from contrived and CML EDTA specimens using Xpert^® BCR-ABL Ultra lysis buffer containing Guanidine Hydrochloride. The frozen contrived blood lysates were stable for 48 hours at RT and 2-8°C and up to 3.5 months at -20°C with ≤2-fold difference from the original testing time-point. The frozen CML blood lysates were stable up to 4 years at -80°C with ≤2-fold difference at ≥ 0.01% (IS) and ≤2.5-fold below 0.01% (IS).

In summary, the presented studies demonstrated that the fresh blood can be stored at RT for 48 hours and at refrigerated temperature for more 3 days without compromising the Xpert^® BCR-ABL Ultra assay sensitive, precision and accuracy. The long term storage can be achieved on DBS at RT for up to 1 year and in PAXgene and frozen lysate at -80^oC for more than 2 years and 4 years, respectively. DBS, the spotted blood method, can potentially benefit the CML patients in resource-poor areas, who can be monitored more frequently during TKI therapy by reducing the sample transport cost significantly.