Shipping blood to a central laboratory in multicenter clinical trials: effect of ambient temperature on specimen temperature, and effects of temperature on mononuclear cell yield, viability and immunologic function

<p>Abstract</p> <p>Background</p> <p>Clinical trials of immunologic therapies provide opportunities to study the cellular and molecular effects of those therapies and may permit identification of biomarkers of response. When the trials are performed at multiple centers, transport and storage of clinical specimens become important variables that may affect lymphocyte viability and function in blood and tissue specimens. The effect of temperature during storage and shipment of peripheral blood on subsequent processing, recovery, and function of lymphocytes is understudied and represents the focus of this study.</p> <p>Methods</p> <p>Peripheral blood samples (n = 285) from patients enrolled in 2 clinical trials of a melanoma vaccine were shipped from clinical centers 250 or 1100 miles to a central laboratory at the sponsoring institution. The yield of peripheral blood mononuclear cells (PBMC) collected before and after cryostorage was correlated with temperatures encountered during shipment. Also, to simulate shipping of whole blood, heparinized blood from healthy donors was collected and stored at 15°C, 22°C, 30°C, or 40°C, for varied intervals before isolation of PBMC. Specimen integrity was assessed by measures of yield, recovery, viability, and function of isolated lymphocytes. Several packaging systems were also evaluated during simulated shipping for the ability to maintain the internal temperature in adverse temperatures over time.</p> <p>Results</p> <p>Blood specimen containers experienced temperatures during shipment ranging from -1 to 35°C. Exposure to temperatures above room temperature (22°C) resulted in greater yields of PBMC. Reduced cell recovery following cryo-preservation as well as decreased viability and immune function were observed in specimens exposed to 15°C or 40°C for greater than 8 hours when compared to storage at 22°C. There was a trend toward improved preservation of blood specimen integrity stored at 30°C prior to processing for all time points tested. Internal temperatures of blood shipping containers were maintained longer in an acceptable range when warm packs were included.</p> <p>Conclusions</p> <p>Blood packages shipped overnight by commercial carrier may encounter extreme seasonal temperatures. Therefore, considerations in the design of shipping containers should include protecting against extreme ambient temperature deviations and maintaining specimen temperature above 22°C or preferably near 30°C.</p

Optimisation and molecular signalling of apoptosis in sequential cryotherapy and chemotherapy combination in human A549 lung cancer xenografts in SCID mice

We define the optimal parameters for combination of cryotherapy (nitrous oxide) with chemotherapy (vinorelbine ditartrate, VNB) treatment and characterise some of the signals involved for apoptosis activation. No advantage appeared when cryotherapy and VNB were combined simultaneously compared to cryosurgery alone. In contrast, tumour volumes were reduced after a sequential treatment schedule, where each individual treatment was separated by 48 h. No significant benefit appeared when the sequential treatment was separated by 24 h, although some individual mice showed a good response. The sequence of treatment had no impact on the observed tumour growth inhibition in mice. The number of apoptotic cells was significantly augmented in the sequential treatment schedule where VNB was administered 48 h before cryotherapy. In this sequential treatment, the number of apoptotic cells correlated with heightened expression of the BH3-only Puma, Noxa and Bim-EL, at both the mRNA and protein levels. No significant change in Bax, Bcl-xL and Bcl-2 mRNA expression was apparent, whereas Mcl-1 expression increased only slightly to a much lower level than BH3-only mRNAs. Our data indicate that 48 h sequential rather than simultaneous cryotherapy with VNB in future cancer cryochemotherapy schedules will enhance the tumour response, and argue that VNB administration, 48 h before cryotherapy, will provoke apoptosis more efficiently