A high-resolution, double-labeling method for the study of in vivo red blood cell aging.

Visual & Circulatory Biophysics Laboratory, Department of Biomedical Engineering, Boston University, MA 02215, USA.
Transfusion (Impact Factor: 3.53). 05/2006; 46(4):578-88. DOI: 10.1111/j.1537-2995.2006.00776.x
Source: PubMed

ABSTRACT Red blood cell (RBC) senescence is a process that has received considerable study, yet remains poorly understood. This has been primarily due to the difficulty in isolating a RBC cohort of narrowly distributed, well-defined age. Biotin labeling has previously been used to produce an identifiable cell cohort of known mean age; however, the variability of RBC age within the cohort is relatively large for most of its existence. Treatments typically employed on animal subjects to reduce this variability can perturb erythropoiesis and result in abnormal RBC aging.
The objective of this study was to improve on the traditional in vivo biotinylation method by introducing a chemically distinct, second labeling step. In this case, digoxigenin was used to label cells 1 to 2 days before the injection of biotin.
It was shown, in the rat, that two identifiable subpopulations of labeled RBCs can be followed over time: a broad, double-labeled cohort and a narrow, single-labeled cohort, the latter consisting of only those cells created between the first and second labeling steps. The utility of this technique was demonstrated by observing the age-dependent exposure of phosphatidylserine in the single-labeled RBCs.
Its capacity to generate a cohort of narrowly distributed age, without the adverse effects associated with animal treatment, should make this a useful method for the study of RBC senescence.

  • [Show abstract] [Hide abstract]
    ABSTRACT: The measurement of red blood cell (RBC) survival has a long history, and a wide variety of methods have been utilized for this purpose. Current methods are of 2 types. First, those that label a representative sample of RBCs of all ages from the blood and then measure their rate of disappearance upon reinfusion. This category includes the (51)Cr and biotin labels. Second, those that use a metabolic precursor or product to determine the turnover of hemoglobin. Examples of these are carbon monoxide production and incorporation of labeled glycine. Recent studies with the covalent, nonradioactive biotin label show its unique suitability for both the accurate measurement of red cell survival and the determination of changes in red cell properties as they age in vivo.
    Transfusion Medicine and Hemotherapy 10/2012; 39(5):302-307. · 1.59 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The goal of this review is to summarize and critically assess information concerning the biotin method to label red blood cells (RBC) for use in studies of RBC and transfusion biology — information that will prove useful to a broad audience of clinicians and scientists. A review of RBC biology, with emphasis on RBC senescence and in vivo survival is included, followed by an analysis of the advantages and disadvantages of biotin labeled RBC (BioRBC) for measuring circulating RBC volume, post-transfusion RBC recovery, RBC lifespan, and RBC age-dependent properties. The advantages of BioRBC over 51Cr RBC labeling, the current reference method, are discussed. Because the biotin method is straightforward and robust, including the ability to follow the entire lifespans of multiple RBC populations concurrently in the same subject, BioRBC offers distinct advantages for studying RBC biology and physiology, particularly RBC survival. The method for biotin labeling, validation of the method, and application of BioRBCs to studies of sickle cell disease, diabetes, and anemia of prematurity are reviewed. Studies documenting the safe use of BioRBC are reviewed; unanswered questions requiring future studies, remaining concerns, and regulatory barriers to broader application of BioRBC including adoption as a new reference method are also presented.
    Transfusion Medicine Reviews 07/2014; · 3.76 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The changes in red blood cells (RBC) as they age and the mechanisms for their eventual removal have been of interest for many years. Proposed age-related changes include dehydration with increased density and decreased size, increased membrane IgG, loss of membrane phospholipid asymmetry, and decreased activity of KCl cotransport. The biotin RBC label allows unambiguous identification of older cells and exploration of their properties as they age. Autologous normal human RBC were labeled ex vivo and, after reinfusion, compared with unlabeled RBC throughout their lifespan. RBC density increased with age, with most of the change in the first weeks. Near the end of their lifespan, RBC had increased surface IgG. However, there was no evidence for elevated external phosphatidylserine (PS) even though older RBC had significantly lower activity of aminophospholipid translocase (APLT). KCl cotransport activity persisted well past the reticulocyte stage, but eventually decreased as the RBC became older. These studies place limitations on the use of density fractionation for the study of older human RBC, and do not support loss of phospholipid asymmetry as a mechanism for human RBC senescence. However, increased levels of IgG were associated with older RBC, and may contribute to their removal from the circulation. Am. J. Hematol., 2012. © 2012 Wiley Periodicals, Inc.
    American Journal of Hematology 09/2012; · 4.00 Impact Factor