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Objective: The currently employed red blood cell reagents have a short shelf life. Some hospitals with a small number of specimens will be unable to utilize them within the validity period, resulting in a substantial increase in the purchase price. Therefore, the method of developing long-term red blood cell reagents is a problem worthy of further study., Methods: In this experiment, the type and concentration of the red blood cell reagent treatment solution were evaluated based on the red blood cell antigen concentration 24 h after treatment. In addition, the qualified glutaraldehyde/paraformaldehyde reagent was stored for six months, and five red blood cell indices were measured every month. At the same time, the detection indices of treated red blood cell reagents and untreated red blood cell reagents were compared., Results: It was discovered that treated red blood cells containing 0.005% GA and 0.05% PFA were more suitable for the preservation of red blood cells than other treated concentrations, and the preservation time could reach six months. The test tube method ( n  = 24) and microcolumn gel card ( n  = 35) were used to determine the accuracy of the treated blood cells containing 0.005% glutaraldehyde +0.05% paraformaldehyde, with an accuracy of 100%., Conclusion: This experiment resulted in the development of a novel reagent for treating red blood cells with glutaraldehyde/paraformaldehyde fixed solution that can effectively prolong its storage time by two to three times that of red blood cell reagents currently on the market.

The mammalian cell nucleus contains different types of membrane-less nuclear bodies (NBs) consisting of proteins and RNAs. Microscopic imaging has been widely applied to study the organization and structure of NBs. However, current fixation methods are not optimized for such imaging: When a fixation method is chosen to maximize the quality of the RNA fluorescence in situ hybridization (FISH), it often limits the labeling efficiency of proteins or affects the ultrastructure of NBs. Here, we report that addition of glyoxal (GO) into the classical paraformaldehyde (PFA) fixation step not only improves FISH signals for RNAs in NBs via augmented permeability of the fixed nucleus and enhanced accessibility of probes, but also largely preserves protein fluorescent signals during fixation and immunostaining. We also show that GO/PFA fixation enables the covisualization of different types of nuclear bodies with minimal impact on their ultrastructures under super-resolution microscopy., (© 2021 Yao et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.)

Mitochondria are highly dynamic organelles, constantly undergoing shape changes, which are controlled by mitochondrial movement, fusion, and fission. Mitochondria play a pivotal role in various cellular processes under physiological and pathological conditions, including metabolism, superoxide generation, calcium homeostasis, and apoptosis. Abnormal mitochondrial morphology and mitochondrial protein expression are always closely related to the health status of cells. Analysis of mitochondrial morphology and mitochondrial protein expression in situ is widely used to reflect the abnormality of cell function in the chemical fixed sample. Paraformaldehyde (PFA), the most commonly used fixative in cellular immunostaining, still has disadvantages, including loss of antigenicity and disruption of morphology during fixation. We tested the effect of ethanol (ETHO), PFA, and glutaraldehyde (GA) fixation on cellular mitochondria. The results showed that 3% PFA and 1.5% GA (PFA-GA) combination reserved mitochondrial morphology better than them alone in situ in cells. Mitochondrial network and protein antigenicity were well maintained, indicated by preserved MitoTracker and mitochondrial immunostaining after PFA-GA fixation. Our results suggest that the PFA-GA combination is a valuable fixative for the study of mitochondria in situ.

Purpose: Chemical fixation is a critical step to retaining cellular targets as naturally as possible. Recent developments in microscopy allow sophisticated detection and measuring techniques with which spatio-temporal molecular alterations are conceivable. In this study, we compare two members of aldehyde fixatives [i.e., glyoxal (Gly) and paraformaldehyde (PFA)] to determine whether Gly, a less toxic dialdehyde fixative that is considered to retain immunoreactivity could provide a successful and consistent cell fixation in favor of PFA in various cell preparations and types., Methods: We document the fixation competence of Gly and PFA side-by-side (with or without Triton X-100 permeabilization) in live- and fixed-cell preparations in mouse oocytes, embryos, and human somatic cells (human umbilical cord-derived mesenchymal stromal cells) using protein quantification by Western blot assay and super-resolution microscopy., Results: Although Gly seemed to act faster than PFA, catastrophic consequences were found not acceptable, especially in oocytes and embryos. Due to cell lysate and immunocytochemistry surveys, it was obvious that PFA is superior to Gly in retaining cellular proteins in situ with little/no background staining. In many samples, PFA revealed more reliable and consistent results regarding the protein quantity and cellular localization corresponding to previously defined patterns in the literature., Conclusion: Although the use of Gly is beneficial as indicated by previous reports, we concluded that it does not meet the requirement for proper fixation, at least for the tested cell types and proteins. However, PFA alone with no addition of TX displayed a significant cytoplasmic loss by generating membrane blebs during fixation.

For safety, designated Select Agents in tissues must be inactivated and viability tested before the tissue undergoes further processing and analysis. In response to the shipping of samples of "inactivated" Bacillus anthracis that inadvertently contained live spores to nonregulated entities and partners worldwide, the Federal Register now mandates in-house validation of inactivation procedures and standardization of viability testing to detect live organisms in samples containing Select Agents that have undergone an inactivation process. We tested and validated formaldehyde and glutaraldehyde inactivation procedures for animal tissues infected with virulent B. anthracis, Burkholderia pseudomallei, Francisella tularensis, and Yersinia pestis. We confirmed that our fixation procedures for tissues containing these Tier 1 Select Agents resulted in complete inactivation and that our validated viability testing methods do not interfere with detection of live organisms. Institutions may use this work as a guide to develop and conduct their own testing to comply with the policy.

Medical knowledge of the skeleton including its structures has improved constantly over the past decades. Advanced imaging methods, mechanical testing and optical techniques have revealed insights into bone architecture and composition. Most of these advancements were possible due to the ex vivo investigation of biological tissues. Investigations of fresh tissue are generally preferred over preserved or fixed samples. However, chemical fixation is sometimes inevitable due to histological procedures or logistical reasons. The aim of this study was to investigate whether short-term chemical fixation with formaldehyde affects bone quality parameters obtained from Raman spectroscopy and if these effects last for intermediate sample storage of several hours. As formaldehyde induces cross-links to the organic components in bone tissue, we hypothesized that collagen-related parameters are particularly affected. Femurs of eight 17-week-old C57BL/6 mice were extracted and divided into two groups (N  =  8  /  group). Samples of the first group were fixed by immersion in 4% formaldehyde (PFA-solution) for 12 h at 4°C (fixed group) while samples of the second group were left untreated (unfixed group). Raman spectroscopy was performed, and repeated after 4 h, to assess whether intermediate storage time influenced the obtained results. Based on resultant spectra, mineral-to-matrix ratio, carbonate-to-phosphate ratio, carbonate-to-amide I ratio, mineral crystallinity and collagen maturity were determined. Carbonate-to-phosphate ratio was the only parameter showing a significant difference between the first and the subsequent measurements. For both groups, ratios showed a decrease in carbonate substitution compared to the first measurement (percentage decrease: 3.1% in fixed, 4.7% in unfixed). Collagen maturity of samples, which were short-term fixed with formaldehyde, was significantly lower than of fresh, unfixed samples (percentage difference: 3.8%). Our study shows that Raman spectroscopy is able to detect changes in collagen structure initiated by formaldehyde and that changes in short-term fixed samples are minimally influencing bone material properties measured with Raman spectroscopy., ((2018) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).)

In a wide range of indoor air pollutants, formaldehyde is one of the most-used and best-known substances. In order to protect human health, many countries have established threshold values for the release of formaldehyde from miscellaneous products and revise them constantly. Compliance with these regulations is usually assessed by emission test chamber measurements or derived methods. To control and improve the mechanisms of an emission test chamber, a reliable reference source with sample mimicking emission properties is required but not available so far. This study describes a permeation-controlled reference source based on the application of paraformaldehyde as formaldehyde releasing polymeric compound. Interactions between the formaldehyde release of the source and the governing chamber parameters temperature, relative humidity and air velocity were investigated in 1 m 3 emission chambers. Depending on the conditions, constant formaldehyde concentrations between approximately 10 ppb and 150 ppb can be adjusted for up to 600 h. A linear correlation between the logarithm of the chamber concentration and the reciprocal temperature was found. The results support the feasibility of the source for validation of emission test chamber performance., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Methylated amines are highly important for a variety of pharmaceutical and agrochemical applications. Existing routes for their formation result in the production of large amounts of waste or require high reaction temperatures, both of which impact the ecological and economical footprint of the methodologies. Herein, we report the ruthenium-catalyzed reductive methylation of a range of aliphatic amines, using paraformaldehyde as both substrate and hydrogen source, in combination with water. This reaction proceeds under mild aqueous reaction conditions. Additionally the use of a secondary phase for catalyst retention and recycling has been investigated with promising results., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Protocols for characterizing cellular phenotypes commonly use chemical fixatives to preserve anatomical features, mechanically stabilize tissue, and stop physiological responses. Formaldehyde, diluted in either phosphate-buffered saline or phosphate buffer, has been widely used in studies of neurons, especially in conjunction with dyes and antibodies. However, previous studies have found that these fixatives induce the formation of bead-like varicosities in the dendrites and axons of brain and spinal cord neurons. We report here that these formaldehyde formulations can induce bead formation in the dendrites and axons of adult rat and rabbit retinal ganglion cells, and that retinal ganglion cells differ from hippocampal, cortical, cerebellar, and spinal cord neurons in that bead formation is not blocked by glutamate receptor antagonists, a voltage-gated Na(+) channel toxin, extracellular Ca(2+) ion exclusion, or temperature shifts. Moreover, we describe a modification of formaldehyde-based fixatives that prevents bead formation in retinal ganglion cells visualized by green fluorescent protein expression and by immunohistochemistry., (© 2014 Wiley Periodicals, Inc.)

Protein-protein interaction at the organelle level can be analyzed by using tagged proteins and assessing Förster resonance energy transfer (FRET) between fluorescent donor and acceptor proteins. Such studies are able to uncover partners in the regulation of proteins and enzymes. However, any organelle movement is an issue for live FRET microscopy, as the observed organelle must not change position during measurement. One of the mobile organelles in plants is the Golgi apparatus following cytoplasmic streaming. It is involved in the decoration of proteins and processing of complex glycan structures for the cell wall. Understanding of these processes is still limited, but evidence is emerging that protein-protein interaction plays a key role in the function of this organelle. In the past, mobile organelles were usually immobilized with paraformaldehyde (PFA) for FRET-based interaction studies. Here, we show that the actin inhibitor Cytochalasin D (CytD) is superior to PFA for immobilization of Golgi stacks in plant cells. Two glycosyltransferases known to interact were tagged with cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP), respectively, coexpressed in Nicotiana benthamiana leaves and analyzed using confocal microscopy and spectral imaging. Fixation with PFA leads to reduced emission intensity when compared to CytD treatment. Furthermore, the calculated FRET efficiency was significantly higher with CytD than with PFA. The documented improvements are beneficial for all methods measuring FRET, where immobilization of the investigated molecules is necessary. It can be expected that FRET measurement in organelles of animal cells will also benefit from the use of inhibitors acting on the cytoskeleton., (Copyright © 2013 International Society for Advancement of Cytometry.)

The SARS-CoV-2 pandemic required the immediate need to transfer inactivated tissue from biosafety level (BSL)-3 to BSL-1 areas to enable downstream analytical methods. No validated SARS-CoV-2 inactivation protocols were available for either formaldehyde (FA)-fixed or glutaraldehyde (GA)-fixed tissues. Therefore, representative tissue from ferrets and hamsters was spiked with 2.2 × 106 tissue culture infectious dose 50% per ml (TCID50/ml) SARS-CoV-2 or were obtained from mice experimentally infected with SARS-CoV-2. SARS-CoV-2 inactivation was demonstrated with 4% FA or 5% GA at room temperature for 72 hours by a titer reduction of up to 103.8 TCID50/ml in different animal tissues with a maximum protein content of 100 µg/mg and a thickness of up to 10 mm for FA and 8 mm for GA. Our protocols can be easily adapted for validating the inactivation of other pathogens to allow for the transfer of biological samples from BSL-3 areas to BSL-1 laboratories. [ABSTRACT FROM AUTHOR]

The condensation/heterocyclization of 2-aminoethanol with formaldehyde and various alcohols afforded 3-(organyloxymethyl)-1,3-oxazolidines. Their yields depended on the nature of the hydroxy compound and increased in parallel with its size. 3,3′-Methylenebis(1,3-oxazolidine) was formed as by-product; it was also synthesized independently by the condensation of 2-aminoethanol with formaldehyde at a ratio of 2:3. Physico-chemical characteristics and 13C NMR spectra of the synthesized compounds are given. NMR signals were assigned using 13C–{1H} heteronuclear double-resonance technique. [ABSTRACT FROM AUTHOR]

The scientific community has responded to the coronavirus disease 2019 (COVID-19) pandemic by rapidly undertaking research to find effective strategies to reduce the burden of this disease. Encouragingly, researchers from a diverse array of fields are collectively working towards this goal. Research with infectious severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is undertaken in high-containment laboratories; however, it is often desirable to work with samples at lower-containment levels. To facilitate the transfer of infectious samples from high-containment laboratories, we have tested methods commonly used to inactivate virus and prepare the sample for additional experiments. Incubation at 80°C, a range of detergents, Trizol reagents, and UV energies were successful at inactivating a high titer of SARS-CoV-2. Methanol and paraformaldehyde incubation of infected cells also inactivated the virus. These protocols can provide a framework for in-house inactivation of SARS-CoV-2 in other laboratories, ensuring the safe use of samples in lower-containment levels. [ABSTRACT FROM AUTHOR]

Lignin is the second most abundant biopolymer and will be an important source for carbon-containing compounds in the future. Based on their similar phenolic structures, lignin has great potential to become a valuable substitute for phenol in phenol-formaldehyde resin adhesives. To meet this aim, the sodium hydroxide (NaOH)-catalyzed reaction of kraft lignin with formaldehyde was studied by using paraformaldehyde (PFA) as a formaldehyde source. The advantage of using PFA, the solid polymer of formaldehyde, is the simple composition of the depolymerized solution. According to the results of differential scanning calorimetry (DSC), the lignin reaction was found to require a high NaOH concentration in order for the reaction with PFA to proceed at reasonably low temperatures compared to the curing temperature of phenol-formaldehyde resins (approximately 150°C). On the other hand, high alkalinity conditions are known to favor the disproportionation of formaldehyde to formic acid and methanol. Due to the moderate reactivity of lignin, the Cannizzaro reaction can compete with the methylolation reaction of lignin. Based on the results of 13C, 31P and 1H-13C heteronuclear single quantum correlation nuclear magnetic resonance (HSQC NMR), methylolation was found to be the main reaction occurring in the lignin-formaldehyde reaction. [ABSTRACT FROM AUTHOR]

The use of Toxavit for pulpal devitalisation in a 30-year-old woman is presented. Improper use of this material resulted in necrosis of soft tissues and bone. The bone sequestrum was removed and examined under an electron microscope, revealing thick accumulation of plaque on the surface of thebone sequestrum and dissolution of bone material under the influence of microorgansims. [ABSTRACT FROM AUTHOR]

Using NMR spectroscopy, the competition between different pathways of the interaction of formaldehyde with n-butylamine, n-octylamine, di-n-butylamine, and di-n-octylamine in aqueous media was studied. The effect of the ratio of secondary amines and formaldehyde on the formation of (N,N-dialkylamino)methanols and N,N,N',N'-tetraalkylmethanediamines has been determined. It has been shown that (N-alkylamino)methanols are formed in high yields when a primary amine is used. The replacement of the butyl radical in the starting amine with the octyl radical has a slight effect on the course of the reaction. [ABSTRACT FROM AUTHOR]

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As a hazardous volatile organic compound (VOC), formaldehyde (FA) is found ubiquitously in both polluted and unpolluted environments. Hence, for the proper control of FA, techniques for rapid and precise quantitation of FA are in great demand. However, because of its low molecular weight and high reactivity, its quantitation generally requires complicated procedures (such as combination of sampling through 2,4-dinitrophenylhydrazine (DNPH) derivatization and detection through high-performance liquid chromatograph (HPLC)). Such an approach, despite proven reliability, is inefficient for fast tracking of FA level under dynamic conditions. To overcome such limitations and to facilitate the near real-time monitoring of gaseous FA, we developed an alternative detection method based on a gas chromatograph/barrier discharge ionization detector interfaced with a large-volume injection system (LVI/GC/BID). The reliability of lab-made gaseous standards of formaldehyde was investigated based on thermal cracking of paraformaldehyde. The developed LVI/GC/BID approach allowed the detection of gaseous FA with a method detection limit (MDL) of about 0.13 ng (0.21 ppm for 0.5 mL sample) over the linear dynamic range of 0.29–5300 ng with relative standard deviation (RSD) of 1.19% compared with the direct injection-based GC/BID MDL of 2.41 ng (6.56 ppm for 0.3 mL sample) and RSD of 3.35%. [ABSTRACT FROM AUTHOR]

In this study, bio-oil from the fast pyrolysis of renewable biomass was used as the raw material to synthesize bio-oil phenol formaldehyde (BPF) resin—a desirable resin for fabricating phenolic-based material. During the synthesis process, paraformaldehyde was used to achieve the requirement of high solid content and low viscosity. The properties of BPF resins were tested. Results indicated that BPF resin with the bio-oil addition of 20% had good performance on oxygen index and bending strength, indicating that adding bio-oil could modify the fire resistance and brittleness of PF resin. The thermal curing behavior and heat resistance of BPF resins were investigated by differential scanning calorimetry (DSC) and thermal gravimetric analysis (TGA). Results showed that adding bio-oil had an impact on curing characteristics and thermal degradation process of PF resin, but the influence was insignificant when the addition was relatively low. The chemical structure and surface characteristics of BPF resins were determined by Fourier transform infrared (FTIR) spectroscopy and scanning electron microscopy (SEM). The analysis demonstrated that adding bio-oil in the amount of 20% was able to improve the crosslinking degree and form more hydrocarbon chains in PF resin. [ABSTRACT FROM AUTHOR]

A validation study for primary formaldehyde gas standards was performed at three National Metrology Institutes: the Korea Research Institute of Standards and Science (KRISS), the National Metrology Institute of Japan (NMIJ) and the National Institute of Metrology of China (NIM). The studied materials had a nominal amount fraction of 2 μmol/mol formaldehyde in nitrogen balance and were prepared in 10-L aluminum cylinders by KRISS. The impurities in the materials were analyzed using a gas chromatograph/atomic emission detector and a Fourier-transform infrared spectrometer (FTIR). The stability of the materials was assessed for 1 year by KRISS using paraformaldehyde as a source for the primary standard gas and a cavity ring-down spectrometer (CRDS) instrument as the measurement method. The amount fraction of formaldehyde in the materials decreased linearly by 0.74 % each month. The studied materials that exhibited similar linear rates of decline were distributed to the participants. After the measurement was completed by the participants, the materials were returned to KRISS and the stability analysis based on the primary standard maintained at KRISS was repeated. NMIJ analyzed the materials using paraformaldehyde as the source of the primary standard of formaldehyde and FTIR analysis, whereas NIM used trioxane as the primary standard gas source and CRDS analysis. The results of the comparison revealed good agreement between the results and were within the expanded uncertainty of 2 % although each of them used different combinations of methods in the generation of primary gas standards and measurements. [ABSTRACT FROM AUTHOR]

The effectiveness of fixatives for fixing biological specimens has long been widely investigated. However, the lowest concentrations of fixatives needed to completely fix whole cells or various cellular structures remain unclear. Using real-time imaging and quantification, we determined the lowest concentrations of glutaraldehyde (0.001-0.005, ~0.005, 0.01-005, 0.01-005, and 0.01-0.1 %) and formaldehyde/paraformaldehyde (0.01-0.05, ~0.05, 0.5-1, 1-1.5, and 0.5-1 %) required to completely fix focal adhesions, cell-surface particles, stress fibers, the cell cortex, and the inner structures of human umbilical vein endothelial cells within 20 min. With prolonged fixation times (>20 min), the concentration of fixative required to completely fix these structures will shift to even lower values. These data may help us understand and optimize fixation protocols and understand the potential effects of the small quantities of endogenously generated aldehydes in human cells. We also determined the lowest concentration of glutaraldehyde (0.5 %) and formaldehyde/paraformaldehyde (2 %) required to induce cell blebbing. We found that the average number and size of the fixation-induced blebs per cell were dependent on both fixative concentration and cell spread area, but were independent of temperature. These data provide important information for understanding cell blebbing, and may help optimize the vesiculation-based technique used to isolate plasma membrane by suggesting ways of controlling the number or size of fixation-induced cell blebs. [ABSTRACT FROM AUTHOR]

Abstract: 3,4-Methylenedioxybenzaldehyde [1-3-benzodioxole-5-carboxaldehyde or heliotropine (HEL)] is widely used in fragrance and pharmaceutical preparations. The synthesis of HEL by Oppenauer''s oxidation of the corresponding 3,4-methylenedioxybenzyl alcohol [piperonyl alcohol (PA)] has been investigated using both homogeneous and heterogeneous catalysts. Commercial paraformaldehyde (PFA) was used as hydrogen acceptor and the oxidation proceeded with high activity and selectivity, operating in liquid phase at 383K, with toluene as solvent and a molar ratio PFA/PA=2.5. In all cases, a critical step was the removal of the possible water coming from the reactants. Homogeneous Al iso-propoxide catalyst gave rise to a very fast and selective reaction, although presenting relevant economic and environmental drawbacks. Among the heterogeneous catalysts, best initial results were obtained using commercial zirconia-type catalysts, with high yield values regardless of catalyst amount and reagent concentration. However, these catalysts cannot be regenerated. Calcined Mg/Al commercial hydrotalcites not only showed good catalytic results (with only lower reaction rates lower than zirconia-type ones), but also an almost complete recovery of the initial activity after calcinations at 823K. [Copyright &y& Elsevier]

MoO3/SiO2 composite with varying amounts of MoO3 loading (1–20 wt.%) were prepared by sol–gel method and calcined at 500 °C. These catalysts were employed for the liquid phase condensation of anisole with paraformaldehyde. All the catalysts were characterized by N2 sorption, XRD, and NH3-TPD. The activities of synthesized MoO3/SiO2 catalysts were compared with p-toluene sulfonic acid ( p-TSA), the most frequently used catalyst for the condensation reactions, and with a supported metal oxide (WO x /ZrO2). Under the similar reaction conditions, synthesized 10 wt.% MoO3/SiO2 catalyst calcined at 500 °C was found to be the most active in the condensation of anisole with paraformaldehyde. [ABSTRACT FROM AUTHOR]

Exposure to aldehydes represents potential risks to human and animal health. Cyclic aldehydes such as benzaldehyde, 2-furaldehyde, and paraldehyde were found to induce formation of stable DNA–protein cross-links (DPXs) in cultured human lymphoma cells. A relationship between increased cytotoxicity and DPX formation was observed with each aldehyde. Paraldehyde is a sedative drug used predominately in treatment of ethanol withdrawal. Paraldehyde was the most potent cross-linking aldehyde studied, yet least cytotoxic. Although DPX formation by aliphatic aldehydes is well-known, this study confirms the potential for cyclic aldehydes to cause formation of DPXs in cultured cells at therapeutically relevant doses. [ABSTRACT FROM AUTHOR]

Summary: This study was designed to analyze and compare tissue preservation efficiency of acetone (AC), formaldehyde (FA) and paraformaldehyde (PFA) on cryosections. Brain, kidney, heart and liver tissue of adult Balb/c mice were fixed with either FA or PFA prior to cryosectioning, or fixed with AC alone immediately after cryosectioning. Hematoxylin and eosin staining showed that AC is a poor fixative in preserving the general tissue and cellular organization. PFA, and to a lesser extent FA, produced significantly better results. Another set of cryosections were further analyzed to test the properties of those fixatives to preserve proteins from specific cell structures. Cytokeratin filaments, F-actin filaments and nuclei were immunolabeled and examined using confocal microscopy. Results demonstrated that, overall, PFA is the best fixative tested. However, FA fixation gave poor results in preserving neuronal tissues. Immunofluorescence confirmed the inefficiency of AC fixation, after which no specific labelling of cytokeratin filaments was detectable. Nevertheless, actin filaments were detectable on AC-fixed samples, a finding that was supported by the quantification of fluorescein-phalloidin binding to F-actin. Overall, the data suggest that AC fixation is unacceptable for preservation of most samples, whereas FA and PFA fixation should be chosen according to the tissues and proteins to be studied. [Copyright &y& Elsevier]

Abstract: Paraformaldehyde fixation of platelets stabilizes surface antigens while altering some that are associated with cellular activation. Present experiments show that the asymmetric distribution of phosphatidylserine in platelets was especially sensitive to paraformaldehyde treatment. It was found that this reagent induced a dose- and time-dependent translocation of phosphatidylserine to the membrane surface as measured by annexin V binding and flow cytometry. The percent phosphatidylserine-positive cells increased from about 5% to >90%. Chelation of extracellular Ca2+ with EGTA partially blocked this translocation. Spectrofluorimetric analysis of fluo-3 loaded platelets indicates that paraformaldehyde caused a concomitant elevation of intracellular Ca2+ concentrations, [Ca2+]i. ATP levels also declined in paraformaldehyde-treated cells, suggesting that the rise in [Ca2+]i ensued in part from decreased activity of calcium pumps. Previous studies indicate that phosphatidylserine externalization arises from Ca2+-activated randomization of membrane phospholipids and decreased transport of phosphatidylserine from the outer to the inner leaflet of the plasma membrane. In light of present results, paraformaldehyde fixation is best avoided particularly in studies involving platelet apoptosis or activation. [Copyright &y& Elsevier]