John A. Widness

Background: In hematologic and transfusion medicine research, measurement of red blood cell (RBC) in vivo kinetics must be safe and accurate. Recent reports indicate use of biotin‐labeled RBC (BioRBC) to determine red cell survival (RCS) offers substantial advantages over ⁵¹Cr and other labeling methods. Occasional induction of BioRBC antibodies has been reported.Study Design and Methods: To investigate the causes and consequences of BioRBC immunization, we reexposed three previously immunized adults to BioRBC and evaluated the safety, antibody emergence, and RCS of BioRBC.Results: BioRBC re‐exposure caused an anamnestic increase of plasma BioRBC antibodies at 5–7 days; all were subclass IgG₁ and neutralized by biotinylated albumin, thus indicating structural specificity for the biotin epitope. Concurrently, specific antibody binding to BioRBC was observed in each subject. As biotin label density increased, the proportion of BioRBC that bound increased antibody also increased; the latter was associated with proportional accelerated removal of BioRBC labeled at density 6 μg/mL. In contrast, only one of three subjects exhibited accelerated removal of BioRBC density 2 μg/mL. No adverse clinical or laboratory events were observed. Among three control subjects who did not develop BioRBC antibodies following initial BioRBC exposure, re‐exposure induced neither antibody emergence nor accelerated BioRBC removal.Discussion: We conclude re‐exposure of immunized subjects to BioRBC can induce anamnestic antibody response that can cause an underestimation of RCS. To minimize chances of antibody induction and underestimation of RCS, we recommend an initial BioRBC exposure volume of ≤10 mL and label densities of ≤18 μg/mL.

Background: Umbilical arterial catheters (UACs) are frequently used in critically ill neonates. UAC are convenient, reliable, and allow for caregiver convenience in performing painless arterial blood sampling. We hypothesized that UAC removal in extremely low birth weight (ELBW) neonates will result in significantly less phlebotomy blood loss (PBL) after correcting for severity of illness. Study Design and Methods: PBL was measured at a single center in 99 ELBW infants who survived to day 28. Individual infant’s paired daily PBL for the two 24-h periods before and after UAC removal were compared using the paired t test. Daily PBL up to 7 days before and 7 days after UAC removal were compared using a logistic regression with mixed model analysis for repeated measures. Cumulative 28-day phlebotomy loss was evaluated by multiple linear regression analysis. Results: PBL 24 h before and after UAC removal were 1.7 mL (95% CI 1.5–1.9) and 0.9 mL (95% CI 0.8–1.0; p < 0.0001), respectively. Cumulative 28-day PBL increased by 2.2 mL (±0.7) per day that a UAC was present with or without correction for severity of illness (p < 0.001). Conclusion: UAC removal is independently associated with a marked decline in PBL. We speculate the ease and convenience of UAC blood sampling lead to more frequent blood testing and greater PBL.

The objective of this review is to provide an integrated historical account of the complex, often convoluted events impacting milk hygiene and its resultant effect on infant mortality from 1875 to 1925. Heat pasteurization of cow's milk is necessary for rendering this important nutrient source safe for humans—particularly infants. Developed by Louis Pasteur in 1864, pasteurization evolved from the commercially important parboiling of wine and beer when the Industrial Revolution was effecting rapid societal change in Western societies. In European and American societies of the early and mid-19th century, infant mortality rates were 30- to 60-fold higher than the current rates of five or six deaths per 1,000 live births per year. With proof of the germ theory of disease came convincing evidence of the role of microbes in the transmission of infections, which led to the discovery that microbial pathogens were transmissible via milk. Diseases caused by milkborne pathogens include human and bovine tub...

The current reference method in the United States for measuring in vivo population red blood cell (RBC) kinetics utilizes chromium‐51 (⁵¹Cr) RBC labeling for determining RBC volume, 24‐hour posttransfusion RBC recovery, and long‐term RBC survival. Here we provide evidence supporting adoption of a method for kinetics that uses the biotin‐labeled RBCs (BioRBCs) as a superior, versatile method for both regulatory and investigational purposes. RBC kinetic analysis using BioRBCs has important methodologic, analytical, and safety advantages over ⁵¹Cr‐labeled RBCs. We critically review recent advances in labeling human RBCs at multiple and progressively lower biotin label densities for concurrent, accurate, and sensitive determination of both autologous and allogeneic RBC population kinetics. BioRBC methods valid for RBC kinetic studies, including successful variations used by the authors, are presented along with pharmacokinetic modeling approaches for the accurate determination of RBC pharmacokinetic variables in health and disease. The advantages and limitations of the BioRBC method—including its capability of determining multiple BioRBC densities simultaneously in the same individual throughout the entire RBC life span—are presented and compared with the ⁵¹Cr method. Finally, potential applications and limitations of kinetic BioRBC determinations are discussed.

BACKGROUND: Critically ill preterm very‐low‐birthweight (VLBW) neonates (birthweight ≤ 1.5 kg) frequently develop anemia that is treated with red blood cell (RBC) transfusions. Although RBCs transfused to adults demonstrate progressive decreases in posttransfusion 24‐hour RBC recovery (PTR₂₄) during storage—to a mean of approximately 85% of the Food and Drug Administration—allowed 42‐day storage—limited data in infants indicate no decrease in PTR₂₄ with storage.STUDY DESIGN AND METHODS: We hypothesized that PTR₂₄ of allogeneic RBCs transfused to anemic VLBW newborns: 1) will be greater than PTR₂₄ of autologous RBCs transfused into healthy adults and 2) will not decrease with increasing storage duration. RBCs were stored at 4°C for not more than 42 days in AS‐3 or AS‐5. PTR₂₄ was determined in 46 VLBW neonates using biotin‐labeled RBCs and in 76 healthy adults using ⁵¹Cr‐labeled RBCs. Linear mixed‐model analysis was used to estimate slopes and intercepts of PTR₂₄ versus duration of RBC storage.RESULTS: For VLBW newborns, the estimated slope of PTR₂₄ versus storage did not decrease with the duration of storage (p = 0.18) while for adults it did (p < 0.0001). These estimated slopes differed significantly in adults compared to newborns (p = 0.04). At the allowed 42‐day storage limit, projected mean neonatal PTR₂₄ was 95.9%; for adults, it was 83.8% (p = 0.0002).CONCLUSIONS: These data provide evidence that storage duration of allogeneic RBCs intended for neonates can be increased without affecting PTR₂₄. This conclusion supports the practice of transfusing RBCs stored up to 42 days for small‐volume neonatal transfusions to limit donor exposure.

BACKGROUND: Biotin‐labeled red blood cells (BioRBCs) are used for in vivo kinetic studies. Because BioRBC dosing occasionally induces antibodies, a sensitive and specific anti‐BioRBC detection assay is needed.STUDY DESIGN AND METHODS: Aims were to 1) develop a gel card assay to evaluate existing, naturally occurring and BioRBC‐induced plasma antibodies, 2) compare gel card and tube agglutination detection results, and 3) test for a relationship of antibody induction and BioRBC dose. Reagent BioRBCs were prepared using sulfo‐NHS biotin ranging from densities 18 (BioRBC‐18) to 1458 (BioRBC‐1458) µg/mL RBCs.RESULTS: Among BioRBC‐exposed subjects, gel card and tube agglutination results were concordant in 21 of 22 adults and all 19 infant plasma samples. Gel card antibody detection sensitivity was more than 10‐fold greater than tube agglutination. Twelve to 16 weeks after BioRBC exposure, induced anti‐antibodies were detected by gel card in three of 26 adults (12%) at reagent densities BioRBC‐256 or less, but in none of 41 infants. Importantly, induced anti‐BioRBC antibodies were associated with higher BioRBC dose (p = 0.008); no antibodies were detected in 18 subjects who received BioRBC doses less than or equal to BioRBC‐18. For noninduced BioRBC antibodies, six of 1125 naïve adults (0.3%) and none of 46 naïve infants demonstrated existing anti‐BioRBC antibodies using reagent BioRBC‐140 or ‐162. Existing anti‐BioRBCs were all neutralized by biotin compounds, while induced antibodies were not.CONCLUSIONS: The gel card assay is more sensitive than the tube agglutination assay. We recommend reagent BioRBC‐256 for identifying anti‐BioRBCs. Use of a low total RBC biotin label dose (≤ BioRBC‐18) may minimize antibody induction.

Background: Reticulocyte hemoglobin content (RET‐He)—an established indicator of iron status in children and adults—was determined in very low birth weight (VLBW) infants.Methods: Longitudinal retrospective RET‐He data in 26 VLBW neonates during the first month of age were compared with: (a) concurrent complete blood counts (CBCs), including hemoglobin (Hb) concentration, reticulocyte count, and immature reticulocyte fraction (IRF), and erythropoietin (EPO) levels; (b) clinical variables; and (c) RET‐He data from the literature for term infants, children, and adults.Results: RET‐He within 24 hr following birth was 31.8 ± 1.1 pg (mean ± SEM). This was followed by an abrupt, significant decline to 28.3 ± 1.1 pg at 2–4 days, and to steady state levels of 28.4 ± 0.5 pg thereafter. The changes in RET‐He were mirrored by changes in plasma EPO, reticulocyte count, and IRF, but not Hb. Steady state RET‐He values after 4 days were significantly lower than RET‐He values for term infants, children, and adults (31.6 ± 0.11, 32.0 ± 0.12, and 33.0 ± 0.13 pg, respectively).Conclusion: Although RET‐He values in VLBW infant were lower than term infants, children, and adults, the significance and mechanism(s) responsible are unknown. The present VLBW infant data are relevant to investigations assessing hemoglobinization following treatment with recombinant human EPO (r‐HuEPO) and/or iron.

In children aged >18 months with cerebral malaria, elevated erythropoietin levels were associated with increased mortality, prolonged coma duration, and a lack of neuroprotection. Caution is warranted in the use of systemic erythropoietin as adjunctive therapy in cerebral malaria. Background. Elevated endogenous plasma erythropoietin (EPO) levels have been associated with protection from acute neurologic deficits in Kenyan children with cerebral malaria (CM). Based on these findings and animal studies, clinical trials of recombinant human EPO (rHuEPO) have been started in children with CM. Recent clinical trials in adults with acute ischemic stroke have demonstrated increased mortality with rHuEPO treatment. We conducted a study in children with CM to assess the relationship of endogenous plasma and cerebrospinal fluid (CSF) EPO levels with mortality and acute and long-term neurologic outcomes. Methods. A total of 210 children between 18 months and 12 years of age with a diagnosis of CM, were enrolled at Mulago Hospital, Kampala, Uganda. Plasma (n = 204) and CSF (n = 147) EPO levels at admission were measured by radioimmunoassay and compared with mortality and neurologic outcomes. Results. After adjustment for age and hemoglobin level, a 1-natural-log increase in plasma EPO level was associated with a 1.74-fold increase in mortality (95% confidence interval, 1.09–2.77, P = .02). Plasma and CSF EPO levels also correlated positively with coma duration (P = .05 and P = .02, respectively). Plasma and CSF EPO levels did not differ in children with vs those without acute or long-term neurologic deficits. Plasma EPO levels correlated positively with markers of endothelial and platelet activation and histidine-rich protein-2 levels, but remained associated with mortality after adjustment for these factors. Conclusions. High endogenous plasma EPO levels are associated with prolonged coma duration and increased mortality in children >18 months of age with CM.