Demonstration of acid and water recovery systems: Applicability and operational challenges in Indian metal finishing SMEs.

Diffusion dialysis, acid retardation and nanofiltration plants were acquired from Europe and demonstrated in several Indian metal finishing companies over a three year period. These companies are primarily small and medium enterprises (SMEs). Free acid recovery rate from spent pickling baths using diffusion dialysis and retardation was in the range of 78-86% and 30-70% respectively. With nanofiltration, 80% recovery rate of rinse water was obtained. The demonstrations created awareness among the metal finishing companies to reuse resources (acid/water) from the effluent streams. However, lack of efficient oil separators, reliable chemical analysis and trained personnel as well as high investment cost limit the application of these technologies. Local manufacturing, plant customization and centralized treatment are likely to encourage the uptake of such technologies in the Indian metal finishing sector.

Relative DNA content in diploid, polyploid, and multiploid species of Paspalum (Poaceae) with relation to reproductive mode and taxonomy.

It is generally accepted that polyploids have downsized basic genomes rather than additive values with respect to their related diploids. Changes in genome size have been reported in correlation with several biological characteristics. About 75 % of around 350 species recognized for Paspalum (Poaceae) are polyploid and most polyploids are apomictic. Multiploid species are common with most of them bearing sexual diploid and apomictic tetraploid or other ploidy levels. DNA content in the embryo and the endosperm was measured by flow cytometry in a seed-by-seed analysis of 47 species including 77 different entities. The relative DNA content of the embryo informed the genome size of the accession while the embryo:endosperm ratio of DNA content revealed its reproductive mode. The genome sizes (2C-value) varied from 0.5 to 6.5 pg and for 29 species were measured for the first time. Flow cytometry provided new information on the reproductive mode for 12 species and one botanical variety and supplied new data for 10 species concerning cytotypes reported for the first time. There was no significant difference between the mean basic genome sizes (1Cx-values) of 32 sexual and 45 apomictic entities. Seventeen entities were diploid and 60 were polyploids with different degrees. There were no clear patterns of changes in 1Cx-values due to polyploidy or reproductive systems, and the existing variations are in concordance with subgeneric taxonomical grouping.

The Retornus-2 study: impact of respiratory muscle training in subacute stroke patients with dysphagia, study protocol of a double-blind randomized controlled trial.

BACKGROUND: Stroke can lead to varying degrees of oropharyngeal dysphagia, respiratory muscle dysfunction and even increase medical complications such as aspiration, malnutrition and death. Recent studies suggest that inspiratory and expiratory respiratory muscle training (IEMT) can improve swallowing efficacy and may reduce aspiration events. The main purpose of this study is to examine whether an 8-week IEMT programme can improve respiratory muscle strength and swallow dysfunction severity in subacute stroke patients with dysphagia. METHODS: Retornus-2 is a two-arm, prospectively registered, randomized controlled study with blinded assessors and the participation of fifty individuals who have suffered a stroke. The intervention group undergoes IEMT training consisting of 5 sets of 10 repetitions, three times a day for 8 weeks. Training loads increase weekly. The control group undergoes a sham-IEMT protocol. The primary outcome examines the efficacy of the IEMT protocol to increase respiratory muscle strength and reduce dysphagia severity. The secondary outcome assesses the longitudinal impact of dysphagia on body composition and nutritional assessment over a 6-month follow-up. DISCUSSION: IEMT induces an improvement in respiratory muscle strength and might be associated with relevant benefits in dysphagia patterns, as well as a reduction in the number of aspiration events confirmed by videofluoroscopy or fiberoptic endoscopic evaluation of swallowing. The description of the impact of swallowing impairment on nutritional status will help develop new strategies to face this known side-effect. TRIAL REGISTRATION: Clinicaltrials.gov NCT03021252. Registered on 10 January 2017. https://clinicaltrials.gov/ct2/results?cond=retornus+2&term=&cntry=ES&state=&city=&dist= WHO trial Registration data set: Due to heavy traffic generated by the COVID-19 outbreak, the ICTRP Search Portal does not respond. The portal recommends other registries such as clinicaltrials.gov. Protocol version: RETORNUS 2_ PROTOCOL_2.

Translation and Validation of the Spanish Version of the SARC-F Questionnaire to Assess Sarcopenia in Older People.

OBJECTIVES: The revised European consensus on sarcopenia definition and diagnosis (EWGSOP2) includes the SARC-F questionnaire, the most valid and consistent sarcopenia screening tool, as the mandatory first step. Our aim was the translation, cross-cultural adaptation, and validation of the SARC-F questionnaire as a culturally-responsive Spanish-language version for the European population. STUDY DESIGN: Cross-sectional descriptive study, applying the two-step WHO methodology for translation and cross-cultural adaptation of health questionnaires, and harmonization with the Mexican-Spanish version. European Union Geriatric Medicine Society recommendations for SARC-F validation in European languages were considered. PARTICIPANTS: Outpatient clinics of a university hospital. INCLUSION CRITERIA: stable, ambulatory (including aids), community-dwelling population ≥65 years old. MAIN OUTCOME MEASURES: The self-reported 5-item SARC-F questionnaire was administered; scores ≥4 indicated sarcopenia. Sensitivity, specificity, accuracy-likelihood ratios, predictive values, and kappa statistics were calculated and consecutively compared with European Working Group on Sarcopenia in Older People (EWGSOP) and EWGSOP2 criteria. RESULTS: This Spanish version, administered in an average 70s, has adequate internal consistency (Cronbach alpha=0.779). For the validation study, 90 (43.3%) of 208 potentially eligible subjects (81.4 ± 5.9 years old, 75.6% women) were included. SARC-F identified 51 (56.7%) subjects with sarcopenia and 39 (43.3%) without the disease. Prevalence was 17.8% per EWGSOP and 25.6% per EWGSOP2 (58% accuracy and fair agreement: sensitivity, 78.3%; specificity, 50.8%). CONCLUSIONS: SARC-F is a feasible tool, suitable for bedside assessment in community-dwelling older patients. Wide diffusion of this culturally-responsible SARC-F Spanish version is expected as EWGSOP2 is adopted and sarcopenia assessment is broadly implemented in Spain.

Failure to achieve a threshold dose of CD34+ CD110+ progenitor cells in the graft predicts delayed platelet engraftment after autologous stem cell transplantation.

In this study, we retrospectively analysed the utility of CD110 expression on CD34(+) cells as a predictor of delayed platelet transfusion independence in 39 patients who underwent autologous peripheral blood stem cell transplantation. Absolute CD34(+) cells and CD34(+) subsets expressing CD110 were enumerated using flow cytometry. Of the 39 patients, 7 required 21 days or more to achieve platelet transfusion independence. Six of the seven patients received a dose of CD34(+)CD110(+) cells below 6.0 x 10(4)/kg while 30 of 32 patients who achieved platelet transfusion independence in <21 days received a dose of CD34(+)CD110(+) cells >6.0 x 10(4)/kg (P<0.001). Patients with delayed platelet engraftment received a median dose of 5.2 x 10(4) CD34(+)CD110(+) cells/kg compared with a median dose of 16.4 x 10(4) cells/kg for those engrafting within 21 days (P=0.003). Further analysis showed that >6.0 x 10(4) CD34(+)CD110(+) cells/kg was highly sensitive (93.8%) and highly specific (85.7%) for achieving platelet transfusion independence within 21 days. Delay in platelet transfusion independence translated into an increased requirement for platelet transfusion (median 6 vs 2 transfusions, P<0.0001). The dose of CD34(+)/CD110(+) cells/kg infused at time of transplantation appears to be an important factor identifying patients at risk of delayed platelet engraftment.

Developmental programming: Interaction between prenatal BPA and postnatal overfeeding on cardiac tissue gene expression in female sheep.

Epidemiologic studies and studies in rodents point to potential risks from developmental exposure to BPA on cardiometabolic diseases. Furthermore, it is becoming increasingly evident that the manifestation and severity of adverse outcomes is the result of interaction between developmental insults and the prevailing environment. Consistent with this premise, recent studies in sheep found prenatal BPA treatment prevented the adverse effects of postnatal obesity in inducing hypertension. The gene networks underlying these complex interactions are not known. mRNA-seq of myocardium was performed on four groups of four female sheep to assess the effects of prenatal BPA exposure, postnatal overfeeding and their interaction on gene transcription, pathway perturbations and functional effects. The effects of prenatal exposure to BPA, postnatal overfeeding, and prenatal BPA with postnatal overfeeding all resulted in transcriptional changes (85-141 significant differentially expressed genes). Although the effects of prenatal BPA and postnatal overfeeding did not involve dysregulation of many of the same genes, they affected a remarkably similar set of biological pathways. Furthermore, an additive or synergistic effect was not found in the combined treatment group, but rather prenatal BPA treatment led to a partial reversal of the effects of overfeeding alone. Many genes previously known to be affected by BPA and involved in obesity, hypertension, or heart disease were altered following these treatments, and AP-1, EGR1, and EGFR were key hubs affected by BPA and/or overfeeding. Environ. Mol. Mutagen. 58:4-18, 2017. © 2016 Wiley Periodicals, Inc.

RET/PTC-induced gene expression in thyroid PCCL3 cells reveals early activation of genes involved in regulation of the immune response.

RET/PTC rearrangements represent key genetic events involved in papillary thyroid carcinoma (PTC) initiation. The aim of the present study was to identify the early changes in gene expression induced by RET/PTC in thyroid cells. For this purpose, microarray analysis was conducted on PCCL3 cells conditionally expressing the RET/PTC3 oncogene. Gene expression profiling 48 h after activation of RET/PTC3 identified a statistically significant modification of expression of 270 genes. Quantitative PCR confirmation of 20 of these demonstrated 90% accuracy of the microarray. Functional clustering of genes with greater than or less than 1.75-fold expression change (86 genes) revealed RET/PTC3-induced regulation of genes with key functions in apoptosis (Ripk3, Tdga), cell-cell signaling (Cdh6, Fn1), cell cycle (Il24), immune and inflammation response (Cxcl10, Scya2, Il6, Gbp2, Oas1, Tap1, RT1Aw2, C2ta, Irf1, Lmp2, Psme2, Prkr), metabolism (Aldob, Ptges, Nd2, Gss, Gstt1), signal transduction (Socs3, Nf1, Jak2, Cpg21, Dusp6, Socs1, Stat1, Stat3, Cish) and transcription (Nr4a1, Junb, Hfh1, Runx1, Foxe1). Genes coding for proteins involved in the immune response and in intracellular signal transduction pathways activated by cytokines and chemokines were strongly represented, indicating a critical role of RET/PTC3 in the early modulation of the immune response.

Recovery of viable CD34+ cells from cryopreserved hemopoietic progenitor cell products.

The number of CD34+ cells infused into patients at the time of autologous or allogeneic transplantation is a clinically important variable, but the viability of these cells has not been extensively documented. In this study, we analyzed the recovery of viable CD34+ cells before and after cryopreservation on 79 autologous stem cell products, using a novel flow cytometry assay without red cell lysis. For 70 PBSC harvest samples, the mean viable CD34+ cell count was 5.98 x 10(6)/kg (range 0.3-23 x 10(6)/kg) before freezing and 5.4 x 10(6)/kg (range 0.2-23 x 10(6)/kg) after thawing. The median recovery was 93% (range 48-107%), with 90% recovery for NHL (range 48-100%, n=34), 83% for multiple myeloma (range 56-106%, n=11), 92.3% for acute leukemia (range 71-100% n=7) and 94.5% for nonhematological malignancies (range 50-107% n=18). Similarly, for autologous bone marrows (n=9) the median recovery of viable CD34+ cells was 90% (range 68-100%). The recovery of viable CD34+ cells for adult (n=51) and pediatric (n=28) stem cell collections was 91 and 94%, respectively. Further examination of the correlation between the kinetics of hematological recovery and the number of viable progenitor cells infused, particularly at the lower end of the accepted dose range, may be warranted.

Geographic heterogeneity of cellular characteristics of acute myeloid leukemia: a comparative study of Australian and Japanese adult cases.

We assessed a large number of adults (368 from Australia and 494 from Japan) with de novo acute myeloid leukemia (AML) to define the biological differences between the two populations. In this study, AML was classified using the French-American-British (FAB) criteria into seven groups (M1-M7). M2 was more common in Japan than in Australia, whereas M4 occurred more frequently in Australia than in Japan. Other FAB subtypes were evenly distributed. Cytogenetically, Japanese M2 displayed a higher frequency of t(8;21) than Australian (33.1% vs 15.3%, P < 0.05). The t(15;17), inv/del(16), 11q23 aberrations and 5/7/8 abnormalities were seen at similar frequencies. Immunophenotypically, Japanese M4/M5 more frequently displayed CD13 and CD14 than Australian, whereas the stem cell markers, CD34 and HLA-DR were observed at a relatively higher rate in Australian M3 than in Japanese M3. The B cell antigen, CD19 was more frequently seen in Japanese M2 than in Australian M2, but found more often in Australian M5 than in Japanese M5. In both populations, a close relationship was observed between the expression of CD19 and t(8;21). These findings suggest different biological characteristics of AML between the two populations, the main differences being generated by a higher frequency of t(8;21) chromosomal abnormality in Japanese AML. Leukemia(2000) 14, 163-168.

Aminopeptidase-N (CD13; gp 150): contrasting patterns of enzymatic activity in blood from patients with myeloid or lymphoid leukemia.

We have investigated the compartmentalization of aminopeptidase-N-like activity in various blood fractions obtained from patients with acute lymphoid (ALL) or myeloid (AML) leukemia. The primary difference appears not to be the absolute level of overall activity, but rather the relative proportions of the different forms of activity detected. Thus, despite similar levels of total aminopeptidase-N-like activity detected in cells from different leukemic groups, true aminopeptidase-N/CD13 activity was only detected in cells derived from AML patients. Even in these patients, however, most of the detected aminopeptidase-N-like activity ( > 80%) could not be attributed to aminopeptidase-N/CD13. In marked contrast, plasma from leukemic patients also contained substantial total aminopeptidase-N-like activity, of which (irrespective of leukemic group) most could be attributed to aminopeptidase-N/CD13. Whilst slightly higher levels of total activity were obtained in plasma from AML patients compared to ALL patients, there was no difference in the relative proportion attributable to aminopeptidase-N/CD13 (approximately 80% of total aminopeptidase-N-like activity). Evaluation of total aminopeptidase-N-like activity present in whole blood gave differential patterns, and whilst only a proportion (20-40% of total aminopeptidase-N-like activity) could be attributed to true aminopeptidase-N/CD13, blood from patients with CD13+ AML showed the greatest activity so attributable. In total, our results outline the complexities of peptidase activities present within blood of leukemic individuals, and may, in part, explain the variability of previous studies attempting to associate prognostic features with phenotypic expression of CD13.

Fifteen cases of t(1;19)(q23;p13.3) identified in an Australian series of 122 children and 80 adults with acute lymphoblastic leukemia.

The t(1;19)(q23;p13) has been reported in up to 6% of cytogenetically abnormal cases of acute lymphoblastic leukaemia (ALL), associated with a pre-B-ALL phenotype. In the 5-year period 1995-1999, we detected t(1;19) in 13 children and 2 adults with newly diagnosed ALL. This represented 10% of pediatric and 2.5% of adult diagnostic ALL samples successfully cultured in one center during this time. There were 9 males and 6 females. The mean age at diagnosis for the 13 children was 6.5 years (range 1.5 to 14 years) and the 2 adults were aged 42 and 45 years. The unbalanced t(1;19) occurred in 7 of 13 children (54%), contrary to the reported excess of unbalanced translocations at 75%; both adults had the unbalanced translocation. At diagnosis, the t(1;19) was the sole abnormality in 4 patients (26%), and in the remainder (74%) was part of a complex karyotype, which included i(7q) (2 patients), hyperdiploidy (2 patients) and del(6q) (2 patients). Correlation of karyotype with white cell, blast and platelet counts, cell surface markers, initial response to chemotherapy and short-term outcome showed no difference between the balanced and unbalanced forms of the translocation in children or whether t(1;19) was present as the sole abnormality or part of a complex karyotype.

The indirect estimation of hemoglobin concentration in whole blood.

The present conventional methods for determination of Hemoglobin (Hb) concentration in whole blood depend on the direct colorimetric measurement of chemically modified Hb following its release from red cells by lysis. This paper examines an alternative, indirect method which does not require initial red cell lysis, corrects for the falsely elevated Hb due to the lipemic plasma and determines whether elevated WBC count effects a change in the Hb value, without the tedious laboratory manipulations currently required to correct for these artifacts. The method is simple, uncomplicated and is based on the observation of a constant ratio (2.98) between the Mean Corpuscular Volume (MCV) and the Mean Corpuscular Hemoglobin (MCH) indices obtained from standard electronic counters in use in most laboratories. The Hb concentration is calculated by the equation: [formula: see text]. The resultant Hb measurements show an acceptable degree of accuracy and precision when compared with the direct measurements obtained from a Coulter Model S + I, even in the presence of a high WBC or lipid.

[Intrauterine malnutrition in rats. II. Study of weight and mortality of the offspring]. / Desnutrição intrauterina em ratos. II Estudo do peso e mortalidade do produto da concepção.

Rats were submitted to protein-calorie deprivation during different periods of gestation and the body weight and mortality of offspring were evaluated at 0, 28 and 90 days of age. The body weight was considered adequate (n) when the values were up to 2 SD below the mean values of control animals, or inadequate (d) when the values were below 2 SD. Rats of the control group (C) were fed ad libitum (protein 21%), and the rats of the experimental groups were fed during all gestational period (D), during the first half (D1) or second half of pregnancy (D2), with a diet containing 1% of protein, and the intake reduced to 50%. Significant differences were observed in the ponderal evolution for all experimental groups. Group D was the most damaged, with the greatest ponderal deficiency and the greatest mortality rates. Similar behavior was observed for D2. Group D1 was similar to C with the best results and the lowest mortality rate.

[Behavior of calcium, phosphorus and magnesium in rats subjected to malnutrition in different stages of the gestational period]. / Comportamento do cálcio, fósforo e magnésio em ratas submetidas á desnutrição durante varias etapas no período gestacional.

An attempt was made to point out the alterations in serum calcium, phosphorus and magnesium, during gestation in female rats fed ad libitum normal diets (protein content, 21%), a protein-calorie deficient diet (protein content, 1%) in a quantity equal to half of that given to normal rats. As the data revealed, in the control group of pregnant rats a significant increase in calcium on the 17th and 19th days was observed, as well as a decrease at the end of gestation. The values remained at higher levels in this group than those detected in the control non-pregnant rats. The above-mentioned increase was not observed in the undernourished pregnant rats, which showed an evident decrease. The behavior of phosphorus in the control pregnant rats was similar to calcium behavior, but this "ion" had not suffered the influence of malnutrition when isolatedly imposed. With regard to magnesium, we noticed that throughout the experiment serum concentration was not influenced either by malnutrition or by gestation. Malnutrition itself was the greatest responsible factor as judged by the differences observed among the groups.

The circadian clock in oral health and diseases.

Most physiological processes in mammals display circadian rhythms that are driven by the endogenous circadian clock. This clock is comprised of a central component located in the hypothalamic suprachiasmatic nucleus and subordinate clocks in peripheral tissues. Circadian rhythms sustain 24-hour oscillations of a large number of master genes controlling the correct timing and synchronization of diverse physiological and metabolic processes within our bodies. This complex regulatory network provides an important communication link between our brain and several peripheral organs and tissues. At the molecular level, circadian oscillations of gene expression are regulated by a family of transcription factors called "clock genes". Dysregulation of clock gene expression results in diverse human pathological conditions, including autoimmune diseases and cancer. There is increasing evidence that the circadian clock affects tooth development, salivary gland and oral epithelium homeostasis, and saliva production. This review summarizes current knowledge of the roles of clock genes in the formation and maintenance of oral tissues, and discusses potential links between "oral clocks" and diseases such as head and neck cancer and Sjögren's syndrome.

A single tube 10-color flow cytometry assay optimizes detection of minimal residual disease in chronic lymphocytic leukemia.

UNLABELLED: Levels of residual disease (RD) are an independent predictor of progression-free survival (PFS) and overall survival (OS) in patients treated for chronic lymphocytic leukemia (CLL). We modified the international standardized approach (ISA) to RD detection using flow cytometry by developing a single tube 10 color antibody assay. METHOD: A single tube incorporated the following monoclonal antibodies: CD81FITC, CD22PE, CD3ECD, CD5PercP5.5, CD20PECY7, CD79bAPC, CD38A700, CD43APC Alexa750, CD19eFluor 450, and CD45KO. A modified ISA gating strategy was developed that removed contaminating events. Sensitivity assays were performed using dilution with normal peripheral blood and bone marrow. Clinical samples were compared using the ISA and the single tube assay. RESULTS: Dilution studies showed that sensitivity of 0.001% was achievable when a minimum of 1.8 × 10(6) total events were acquired. One hundred twenty-nine samples were analyzed and showed RD levels from 0.003 to 22%. In 80 samples analyzed with both assays, there was an excellent correlation between the two methods (slope = 1.0, intercept = 0.07 and R2 = 0.992) and results from Bland-Altman analysis showed a bias of 0.04 ± 0.38 with 95% confidence interval of -0.71 to 0.79. Removal of contaminating events in the single tube assay led to a significant reduction in RD values (P = 0.0014). CONCLUSION: The single tube 10-color assay for the detection of RD in CLL provides equivalent results to the ISA but requires fewer cells, uses fewer reagents, and allows for simpler analysis. By directly removing contaminating events, it improves the accuracy of CLL RD detection and may reclassify the status of some patients following chemotherapy.

Progression of BRAF-induced thyroid cancer is associated with epithelial-mesenchymal transition requiring concomitant MAP kinase and TGFß signaling.

Mice with thyroid-specific expression of oncogenic BRAF (Tg-Braf) develop papillary thyroid cancers (PTCs) that are locally invasive and have well-defined foci of poorly differentiated thyroid carcinoma (PDTC). To investigate the PTC-PDTC progression, we performed a microarray analysis using RNA from paired samples of PDTC and PTC collected from the same animals by laser capture microdissection. Analysis of eight paired samples revealed a profound deregulation of genes involved in cell adhesion and intracellular junctions, with changes consistent with an epithelial-mesenchymal transition (EMT). This was confirmed by immunohistochemistry, as vimentin expression was increased and E-cadherin lost in PDTC compared with adjacent PTC. Moreover, PDTC stained positively for phospho-Smad2, suggesting a role for transforming growth factor (TGF)ß in mediating this process. Accordingly, TGFß-induced EMT in primary cultures of thyroid cells from Tg-Braf mice, whereas wild-type thyroid cells retained their epithelial features. TGFß-induced Smad2 phosphorylation, transcriptional activity and induction of EMT required mitogen-activated protein kinase (MAPK) pathway activation in Tg-Braf thyrocytes. Hence, tumor initiation by oncogenic BRAF renders thyroid cells susceptible to TGFß-induced EMT, through a MAPK-dependent process.

Comparison of rat olfactory mucosal responses to carcinogenic and non-carcinogenic chloracetanilides.

Alachlor and butachlor are chloracetanilide herbicides that induce olfactory tumors in rats, whereas propachlor does not. The mechanism by which alachlor induces tumors is distinct from many other nasal carcinogens, in that alachlor induces a gradual de-differentiation of the olfactory mucosa (OM) to a more respiratory-like epithelium, in contrast to other agents that induce cytotoxicity, followed by an aberrant regenerative response. We studied biochemical and genomic effects of these compounds to identify processes that occur in common between alachlor- and butachlor-treated rats. Because we have previously shown that matrix metalloproteinase-2 (MMP2) is activated in OM by alachlor, in the present studies we evaluated both MMP2 activation and changes in OM gene expression in response to carcinogenic and non-carcinogenic chloracetanilide treatments. All three chloracetanilides activated MMP2, and >300 genes were significantly up- or downregulated between control and alachlor-treated rats. The most significantly regulated gene was vomeromodulin, which was dramatically upregulated by alachlor and butachlor treatment (>60-fold), but not by propachlor treatment. Except for similar gene responses in alachlor- and butachlor-treated rats, we did not identify clear-cut differences that would predict OM carcinogenicity in this study.

Fresh PBSC harvests, but not BM, show temperature-related loss of CD34 viability during storage and transport.

BACKGROUND: The optimum conditions for storage and transport of freshly harvested HPC in the liquid state are uncertain. It is not specified in commonly applied standards for stem cell transplantation. We used a viable CD34 assay to determine the optimum temperature for maintaining progenitor cell viability in freshly harvested BM and PBSC. Our aim was to identify standardized conditions for storage and transport of marrow or peripheral blood products that would optimize CD34 recovery, leading to better transplant outcomes. METHODS: Samples were aseptically removed from 46 fresh HPC harvests (34 PBSC and 12 BM) and stored at refrigerated temperature (2-8 degrees C), room temperature (18-24 degrees C) and 37 degrees C for up to 72 h. Samples were analyzed for viable CD34+ cells/microL at 0, 24, 48 and 72 h. RESULTS: The mean viable CD34+ yield prior to storage was 7.7 x 10(6)/kg (range 0.7-30.3). The mean loss of viable CD34+ cells in HPC products at refrigerated temperature was 9.4%, 19.4% and 28% at 24, 48 and 72 h, respectively. In contrast, the mean loss of viable CD34+ cells at room temperature was 21.9%, 30.7% and 43.3% at 24, 48 and 72 h, respectively. No viable CD34+ cells remained after storage at 37 degrees C for 24 h. Only PBSC products and not BM showed temperature-related loss of CD34 viability. Greater loss of viable CD34+ cells was observed for allogeneic PBSC compared with autologous PBSC. DISCUSSION: These results demonstrate that the optimum temperature for maintaining the viability of CD34+ cells, during overnight storage and transport of freshly harvested HPC, is 2-8 degrees C. These findings will allow the development of standard guidelines for HPC storage and transport.

Detection of intracellular lymphoid differentiation antigens by flow cytometry in acute lymphoblastic leukemia.

The value of flow cytometric detection of the intracellular lymphoid differentiation antigens CD3 and CD22 in the differential diagnosis of acute leukemia was assessed in cases of acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), and leukemic cell lines. Cells were fixed in 0.25% paraformaldehyde at 4 degrees C for 60 min, permeabilized with 0.2% Tween 20 at 37 degrees C for 15 min, then stained with CD3 or CD22 monoclonal antibodies by indirect immunofluorescence. Cytoplasmic CD22 was detected on greater than 20% (mean 55%; range 20-87%) of blasts from all 20 cases of precursor-B ALL analyzed. The percentage of cells with cytoplasmic CD22 was greater than that with membrane CD22 in all except 2 cases of precursor-B ALL. Cytoplasmic CD22 was not detected in 8 cases of precursor-T ALL, 4 T-leukemia cell lines, or in 7 cases of AML. In contrast, cytoplasmic CD3 was detectable by flow cytometry in all 8 cases of precursor-T ALL, but not in precursor-B ALL, pre-B leukemia cell lines, or in AML. These results confirm that cytoplasmic CD3 and CD22 are excellent markers of the early T and B lineages in ALL and can be reliably detected by flow cytometry. This technique should be a valuable addition to routine immunophenotyping for classification of acute leukemia.