Influence of Standard Laboratory Procedures on Measures of Erythrocyte Damage

Wiegmann, Lena; Zélicourt, Diane de; Speer, Oliver; Muller, Alissa; Goede, Jeroen S.; Seifert, Burkhardt; Kurtcuoglu, Vartan

doi:10.3389/fphys.2017.00731

Cited by 18 publications

(27 citation statements)

References 35 publications

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“…When RBCs are pipetted into a glutaraldehyde solution they are believed to be fixed within 1 s (Sutera and Mehrjardi, 1975). This means that if the pipetting creates a considerable flow (compare also Wiegmann et al, 2017), the cellular shape adaptation to flow (Lanotte et al, 2016; Quint et al, 2017) is also conserved, which is a deviation from RBC shape in stasis (Figure 5) and must be regarded as an artifact. The cells marked by red circles in Figure 5 include knizocytes (trilobes), a clear indication for flow induced cell shape changes (Lanotte et al, 2016).…”

Section: Resultsmentioning

confidence: 99%

Glutaraldehyde – A Subtle Tool in the Investigation of Healthy and Pathologic Red Blood Cells

et al. 2019

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Glutaraldehyde is a well-known substance used in biomedical research to fix cells. Since hemolytic anemias are often associated with red blood cell shape changes deviating from the biconcave disk shape, conservation of these shapes for imaging in general and 3D-imaging in particular, like confocal microscopy, scanning electron microscopy or scanning probe microscopy is a common desire. Along with the fixation comes an increase in the stiffness of the cells. In the context of red blood cells this increased rigidity is often used to mimic malaria infected red blood cells because they are also stiffer than healthy red blood cells. However, the use of glutaraldehyde is associated with numerous pitfalls: (i) while the increase in rigidity by an application of increasing concentrations of glutaraldehyde is an analog process, the fixation is a rather digital event (all or none); (ii) addition of glutaraldehyde massively changes osmolality in a concentration dependent manner and hence cell shapes can be distorted; (iii) glutaraldehyde batches differ in their properties especially in the ratio of monomers and polymers; (iv) handling pitfalls, like inducing shear artifacts of red blood cell shapes or cell density changes that needs to be considered, e.g., when working with cells in flow; (v) staining glutaraldehyde treated red blood cells need different approaches compared to living cells, for instance, because glutaraldehyde itself induces a strong fluorescence. Within this paper we provide documentation about the subtle use of glutaraldehyde on healthy and pathologic red blood cells and how to deal with or circumvent pitfalls.

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Section: Resultsmentioning

confidence: 99%

Glutaraldehyde – A Subtle Tool in the Investigation of Healthy and Pathologic Red Blood Cells

et al. 2019

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“…High speed centrifugation with or without contact with Percoll used for RBC fractionation was shown to impact RBC properties by causing swelling and the corresponding shift in O_min and O_hyper ( Figures 2Aa , Ab ; Wiegmann et al, 2017 ). While we did not see a decrease of EI_max ( Figure 2Ac ), the others have reported a drop in EI_max after centrifugation ( Nemeth et al, 2015 ; Kiss et al, 2016 ; Wiegmann et al, 2017 ). These observations make us question the feasibility of translation of the properties reported for the age-matched RBC after fractionation to the in vivo conditions.…”

Section: Discussionmentioning

confidence: 99%

Donor Age and Red Cell Age Contribute to the Variance in Lorrca Indices in Healthy Donors for Next Generation Ektacytometry: A Pilot Study

et al. 2021

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The ability of red blood cells (RBCs) to transport gases, their lifespan as well as their rheological properties invariably depend on the deformability, hydration, and membrane stability of these cells, which can be measured by Laser optical rotational red cell analyser (Lorrca® Maxsis, RR Mechatronics). The osmoscan mode of Lorrca is currently used in diagnosis of rare anemias in clinical laboratories. However, a broad range of normal values for healthy subjects reduces the sensitivity of this method for diagnosis of mild disease phenotype. In this pilot study, we explored the impact of age and gender of 45 healthy donors, as well as RBC age on the Lorrca indices. Whereas gender did not affect the Lorrca indices in our study, the age donors had a profound effect on the O_hyper parameter. To study the impact of RBC age on the osmoscan parameters, we have isolated low (L)-, medium (M)-, or high (H)- density fractions enriched with young, mature, and senescent RBCs, respectively, and evaluated the influence of RBC age-related properties, such as density, morphology, and redox state, on the osmoscan indices. As before, O_hyper was the most sensitive parameter, dropping markedly with an increase in RBC density and age. Senescence was associated with a decrease in deformability (EI_max) and tolerability to low and high osmolatites (Area). L-fraction was enriched with reticulocytes and cells with high projected area and EMA staining, but also contained a small number of cells small in projected area and most likely, terminally senescent. L-fraction was on average slightly less deformable than mature cells. The cells from the L-fraction produced more oxidants and NO than all other fractions. However, RBCs from the L-fraction contained maximal levels of reduced thiols compared to other fractions. Our study suggests that reference values for O_hyper should be age-stratified, and, most probably, corrected for the average RBC age. Further multi-center study is required to validate these suggestions before implementing them into clinical practice.

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“…Free haemoglobin concentration (g L À1 ) measured for either 5 or 10 min showed the same result for control and centrifuged sample. 21 (e) Impact of number of centrifugation cycles on microRNAs. 38 Higher concentration of serum was observed at second centrifugation than the first cycle.…”

Section: Operating Modesmentioning

confidence: 99%

“…18 The apheresis procedure remains challenging due to the complexity of blood matrix, the variations in blood composition between donors 19 and risks of inducing cell damage. 20 Apheresis techniques can affect cell integrity and biological function 21,22 due to the shear stress experienced during the extraction processes. In addition, only a limited number of different components can be collected simultaneously within a single apheresis procedure due to long treatment times which can lead to significant risks of blood contamination caused by transfusion-transmitted pathogens such as HIV, hepatitis B and C virus, prions, to name a few.…”

Section: Introductionmentioning

confidence: 99%

Blood apheresis technologies – a critical review on challenges towards efficient blood separation and treatment

et al. 2021

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Influence of Standard Laboratory Procedures on Measures of Erythrocyte Damage

Cited by 18 publications

References 35 publications

Glutaraldehyde – A Subtle Tool in the Investigation of Healthy and Pathologic Red Blood Cells

Glutaraldehyde – A Subtle Tool in the Investigation of Healthy and Pathologic Red Blood Cells

Donor Age and Red Cell Age Contribute to the Variance in Lorrca Indices in Healthy Donors for Next Generation Ektacytometry: A Pilot Study

Blood apheresis technologies – a critical review on challenges towards efficient blood separation and treatment

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