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1.
R Fayer  T Nerad 《Applied microbiology》1996,62(4):1431-1433
Microcentrifuge tubes containing 8 x 10(6) purified oocysts of Cryptosporidium parvum suspended in 400 microliters of deionized water were stored at 5 degrees C for 168 h or frozen at -10, -15, -20, and -70 degrees C for 1 h to 168 h and then thawed at room temperature (21 degrees C). Fifty microliters containing 10(6) oocysts was administered to each of five to seven neonatal BALB/c mice by gastric intubation. Segments of ileum, cecum, and colon were taken for histology from each mouse 72 or 96 h later. Freeze-thawed oocysts were considered viable and infectious only when developmental-stage C. parvum organisms were found microscopically in the tissue sections. Developmental-stage parasites were not found in tissues from any mice that received oocysts frozen at -70 degrees C for 1, 8, or 24 h. All mice that received oocysts frozen at -20 degrees C for 1, 3, and 5 h had developmental-stage C. parvum; one of 6 mice that received oocysts frozen at -20 degrees C for 8 h had a few developmental-stage parasites; mice that received oocysts frozen at -20 degrees C for 24 and 168 h had no parasites. All mice that received oocysts frozen at -15 degrees C for 8 and 24 h had developmental-stage parasites; mice that received oocysts frozen at -15 degrees C for 168 h had no parasites. All mice that received oocysts frozen at -10 degrees C for 8, 24, and 168 h and those that received oocysts stored at 5 degrees C for 168 h had developmental-stage parasites. These findings demonstrate for the first time that oocysts of C. parvum in water can retain viability and infectivity after freezing and that oocysts survive longer at higher freezing temperatures.  相似文献   

2.
Long-time storage of faecal samples is necessary for investigations of intestinal microfloras. The aim of the present study was to evaluate how the viability and the composition of the Escherichia coli flora are affected in faecal samples during different storage conditions. Four fresh faecal samples (two from calves and two from infants) were divided into sub-samples and stored in four different ways: with and without addition of glycerol broth at -20 degrees C and at -70 degrees C. The viability and the phenotypic diversity of the E. coli flora in the sub-samples were evaluated after repeated thawings and after storage during 1 year. The samples stored for 1 year without thawing were also kept at room temperature for 5 days and subsequently analysed. According to phenotyping (PhP analysis) of 32 isolates per sample on day 0, all four samples contained two dominating strains of E. coli each, and between one and eight less common strains. Samples that were stored at -70 degrees C in glycerol broth showed equal or even higher bacterial numbers as the original samples, even after repeated thawings, whereas samples stored at -20 degrees C showed a considerably lower survival rate, also with addition of glycerol. Sub-samples containing glycerol broth that were kept at room temperature after storage for 1 year showed a clear increase in the number of viable cells as well as in diversity. The diversities in each sub-sample showed a tendency to decrease after several thawings as well as after storage. Generally, the E. coli populations in samples stored at -20 degrees C were less similar to the population of the original sample than that in samples stored at -70 degrees C. Samples that had been mixed with glycerol broth had an E. coli flora more similar to that in the original sample than those without glycerol broth. Furthermore, the sub-samples that were kept at room temperature after storage for 1 year generally were more similar to the original samples than if they were processed directly. We conclude that for long time storage of faecal samples, storage at -70 degrees C is preferable. If samples have to be thawed repeatedly, addition of glycerol is preferable both for samples stored at -70 degrees C and for samples stored at -20 degrees C. Our data also have indicated that when E. coli isolates from faecal samples are selected for, e.g. analysis of virulence factors, it is necessary to pick several isolates per sample in order to obtain at least one isolate representing the dominating strain(s).  相似文献   

3.
In the present investigation the storage effect of AcD-AG and CPD-AG-stabilizers on thrombocytes was tested. The platelets were stored in platelet-rich plasma (PRP) at 4 degrees C or room temperature for 3 days. The concentrates gained by it were marked with Na251CrO4 and reinjected. The thrombocytokinetic parameters were evaluated. The results show that storage with the help of the mentioned stabilizers can be made to a certain extent only. Platelets stored in AcD-AG stabilizerhad a survival time of 2.7 +/- 1.1 days towards 9.0 +/- 1.0 days of fresh whole blood concentrates. The survival time of CPD-AG thrombocytes stored at 4 degrees C for 3 days amounted to 2.0 +/- 0.5 days. Storage of CPD-AG platelets at room temperature showed favourable results. Their survival time amounted to 6.2 +/- 0.6 days. Measurements of surface activity above the spleen and the liver indicate that degradation of stored platelets is mainly performed in the spleen. Problems of liquids storing in view of the significance of therapeutic thrombocyte substitution for hospitals are referred to.  相似文献   

4.
目的:探讨重组人尿激酶原(rhPro-UK)冻干产品的稳定性。方法:采用S-2444发色底物法测定贮存在4℃和-20℃的rhPro-UK冻干产品的活性、单链比例随时间的变化规律;用SDS-PAGE及RP-HPLC肽图分析贮存在-20℃的rhPro-UK冻干产品的结构与组成的变化。结果:4℃保存3年后的rhPro-UK冻干产品的总活性和单链比例基本没有变化,但随着贮存时间的延长,有部分产品降解,如贮存78个月的样品,总活性可降低13%~15%;-20℃保存78个月后,rhPro-UK冻干产品的总活性和单链比例未见明显变化。SDS-PAGE及RP-HPLC肽图图谱显示,-20℃贮存78个月后的rhPro-UK冻干产品的组成和结构没有变化。结论:rhPro-UK冻干产品在4℃的贮存寿命可达3年;长期贮存于-20℃的rhPro-UK冻干产品,其总活性、单链比例及结构组成非常稳定。  相似文献   

5.
1. The effect of storage conditions of rainbow trout (Salmo gairdneri) liver on monooxygenase and conjugation enzyme activities was studied. Fish livers or whole fish were frozen and stored for various periods of time at -4, -20 or -80 degrees C. 2. Freezing the whole fish at -20 degrees C affected the biotransformation enzyme activities dramatically. The loss of monooxygenase activity exceeded up to one-tenth of the initial rate in 17 days. UDP-Glucuronosyltransferase activity increased 50%. Glutathione S-transferase appeared to be the most durable enzyme. 3. When the whole fish were stored in an ice-bath at -4 degrees C for up to 24 hr the activities measured decreased only half of that when frozen for 3 days. 4. When it is impossible to freeze the tissues studied in liquid nitrogen the activities are best preserved when whole, decapitated, bled fish are kept in an ice-bath for less than 24 hr.  相似文献   

6.
T Kojima  T Soma  N Oguri 《Cryobiology》1987,24(3):247-255
The aim of the present study was to examine the effects of various conditions of addition and dilution of dimethyl sulfoxide (Me2SO) and 37 degrees C equilibration, and also the effects of freezing in the solution which was prepared in advance and stored in plastic straws at -20 degrees C on the viability of rabbit morulae thawed rapidly. The embryos were cooled from room temperature to -30 degrees C at 1 degree C/min in the presence of 1.5 M Me2SO using a programmable liquid nitrogen vapor freezing machine with an automatic seeding device, then cooled rapidly, and stored in liquid nitrogen. The frozen straws were thawed rapidly (greater than 1000 degrees C/min). When Me2SO was added in a single step, equilibrated with embryos at 37 degrees C for 15 min and diluted out in a single step, a very high survival was obtained: transferable/recovered, 90%: developed/recovered, 96%. When embryos were pipetted into 1.5 M Me2SO that was prepared in advance, stocked in straws at -20 degrees C, and cooled, the proportions of transferable and developed embryos were equivalent to those of embryos frozen in the solution that was prepared immediately before use.  相似文献   

7.
Hodge G  Markus C  Nairn J  Hodge S 《Cytokine》2005,32(1):7-11
Intracytoplasmic detection of leucocyte cytokines has become a powerful tool for the characterisation of cytokine-producing cells in heterogeneous cell populations, however the effect of specimen storage conditions is unknown. The aim of this study was to determine the effect of whole blood stored at room temperature (RT) or 4 degrees C, on intracellular cytokine production by T cells and monocytes. In cell cultures stored at RT or 4 degrees C for 24h, significant changes in several leucocyte cytokines/chemokines were shown compared to blood cultures stimulated at time=0. There was a significant decrease in IL-2, IL-4 and TNFalpha production by CD4+ T cells in blood cultures stored at RT but an increase in IL-2 in cultures at 4 degrees C. There was a significant decrease in TGFbeta production by CD4+ and CD8+ T cells in cultures kept at RT or 4 degrees C. There was a significant increase in MCP-1 and MCP-3 production by monocytes in blood cultures kept at RT or 4 degrees C. There was a decrease in IL-12 production by monocytes in cultures kept at 4 degrees C, whereas IL-10 production was decreased at RT and increased in cultures kept at 4 degrees C. Blood stored at 4 degrees C showed less immunomodulatory changes than blood kept at RT although overall a possible Th1 bias at 4 degrees C.  相似文献   

8.
Unpasteurized skim milk was storaged in a frozen state at ?7°C or ?20°C for up to several months. There was no increase of non casein and non protein nitrogens, but a slight increase of free tyrosine and a slight decrease of alkaline phosphatase activity were detected when storage period was prolonged. Destabilization occurred solely in caseinate complex, but non micellar casein appeared to be stable.

The contents of calcium and inorganic phosphate in the caseinate complex separated by ultracentrifugation were increased appreciably after frozen storage. The viscosity characteristics of frozen storaged skim milk was also investigated.

Caseinate complex was ultracentrifugally separated from skim milk before and after frozen storage, and then lyophilized. Skim milk itself was also lyophilized before and after frozen storage. Dispersibility was examined on the reconstituted suspension of the lyophilized samples.

The lyophilized sample from frozen storaged milk was much less dispersible than the lyophilized control sample prepared before frozen storage. However, when lyophilized samples were once resolved with reagents such as urea and potassium oxalate and then dialyzed against fresh milk, stable micelle resulted in both samples prepared before and after frozen storage.

Some reduction of dispersibility occurred during lyophilization and subsequent storage in a dried state in the caseinate complex prepared before frozen storage. This reduction was small when skim milk was lyophilized and stored.  相似文献   

9.
When equal volumes of 6% lactose and a broth culture of Yersinia pestis were mixed before freezing, approximately 50% of the cells survived lyophilization and reconstitution on the following day. Concomitantly, the number of viable cells per 50% lethal dose increased from about 16 to 125 organisms. On subsequent storage of the lyophilized cells under vacuum in glass ampoules at 4 degrees C for 25 years, more than 25% of the cells remained viable. When stored cultures were assayed immediately after reconstitution, virulence for mice was significantly reduced (as many as 4,000 cells/50% lethal dose), but the virulence was fully restored when reconstituted cultures were held for 24 h at room temperature, or when a subculture was prepared in fresh medium.  相似文献   

10.
AIMS: To obtain preliminary data on the microbiology and hurdles to pathogen growth in the traditional Pacific Island food, povi masima, which is essentially beef brisket cured in brine. METHODS AND RESULTS: Six containers of povi masima were prepared and two were inoculated with five enterotoxigenic strains of Staphyloccocus aureus. The povi masima were divided into two lots each containing two uninoculated control and an inoculated container. Lot 1 was incubated at room temperature (20 degrees C) and lot 2 under refrigeration (4-5 degrees C) for up to 98 days. During storage, samples were removed and tested for aerobic plate count, coagulase-producing Staphylococci, Clostridium perfringens, staphylococcal enterotoxin and various chemical parameters of the food. Coagulase-producing Staphylococci and aerobic plate counts grew to high levels in both the inoculated and uninoculated lots stored at room temperature, but enterotoxin was only detected at one time point in these lots and this may represent a false positive result. The concentration of NaCl in the meat increased with time as concentrations equilibrated, and nitrite was rapidly lost in those lots stored at room temperature. Storage at 4-5 degrees C prevented proliferation of coagulase-producing Staphylococci. CONCLUSIONS: For safe curing and storage, this food should be kept under refrigeration as this prevented growth of staphylococci. Optimum storage would also be achieved with improved attempts to ensure equal distribution of NaCl prior to storage. SIGNIFICANCE AND IMPACT OF THE STUDY: Under conditions traditionally used to cure and store this food, enterotoxigenic staphylococci can grow to numbers where toxigenesis might occur, especially during the early stages of curing where the salt has not diffused from the brine into the meat.  相似文献   

11.
Stability of aminoglycoside phosphotransferases, adenylyltransferases and acetyltransferases isolated from various sources was studied. The enzymes were characterized by different substrate profiles. They were stored at a temperature of -10 degrees C in the form of frozen solutions or at a temperature of 4 degrees C in the lyophilized form. It was shown that lyophilization markedly increased the stability of the enzymes inactivating aminoglycoside antibiotics. Aminoglycoside phosphotransferases and adenylyltransferases with streptomycin as substrate were less stable than aminoglycoside phosphotransferases with neomycin as substrate. Aminoglycoside acetyltransferases from Streptomyces fradiae 918 producing neomycin were least stable among the enzymes studied. Lyophilized enzymes as a possible stabilizer of ATP added to the preparations had no significant effect on their stability during storage.  相似文献   

12.
A method is described for storage of chromatin and cytoplasm for electrophoresis. Chromatin was prepared from isolated nuclei, dialyzed against 10?4m phenylmethylsulfonyl fluoride and 0.002 m EDTA, and stored as a lyophilized powder for a period of 4 weeks. Polyacrylamide slab-gel electrophoresis of chromatin treated in this manner showed improved resolution and considerably less degradation than chromatin samples solubilized in sodium dodecyl sulfate and frozen at ?20°C in liquid N2 or left at room temperature. Electrophoresis of stored cytoplasmic proteins gave similar results. This method allows convenient concentration of the sample. It also avoids the use of detergents and gives flexibility in the choice of buffers for later electrophoresis.  相似文献   

13.
Parameters affecting the yield of DNA from human blood   总被引:34,自引:0,他引:34  
An examination of variables affecting the yield of DNA from blood was undertaken in order to improve sample processing and to evaluate alternative methods of mailing blood samples for DNA analysis. A rapid, high-yield method was developed for the isolation of high-molecular-weight DNA from fresh and frozen blood. In addition, the following observations were made: (1) Of the anticoagulants examined, acid citrate dextrose (ACD) solution B was found to be superior to EDTA and heparin for preserving yields of DNA during incubation at room temperature. If DNA is isolated from frozen blood, high yields of undegraded DNA are achieved after incubation at 23 degrees C for 5 days with ACD solution B. (2) High yields of undegraded DNA are obtained from blood stored with ACD solution B for at least 1 day at 42 degrees C, 5 days at 0 degrees C, or 1 month at -20 degrees C. (3) Three cycles of freezing and thawing may have little if any affect on the yield of DNA. The results indicate that blood for DNA extraction may be mailed in an ambient temperature container and, in many cases, sent by first-class mail rather than by overnight delivery services.  相似文献   

14.
This study was designed to examine whether rat spermatozoa after freeze-drying and 1-year storage can participate in full-term development following intracytoplasmic sperm injection (ICSI). Cauda epididymal spermatozoa from Crlj:Wistar rats were frozen in liquid nitrogen (LN(2)), first dried for 14 hr at 0.37 hPa and then for 3 hr at 0.001 hPa. The dried spermatozoa were stored for 1 year in a desiccator at +25 degrees C, or in a refrigerator at +4 degrees C, or in LN(2) at -196 degrees C. Controls consisted of sperm that had only been frozen and stored in LN(2). After being stored, spermatozoa were sonicated to dissociate the sperm tail and were injected into oocytes from superovulated Slc:SD rats. The respective fertilization rates of oocytes injected with frozen sperm, or with freeze-dried sperm stored at +25, +4, and -196 degrees C were 79%, 75%, 70%, and 73%. However, the corresponding cleavage rates of injected oocytes were 63%, 1%, 38%, and 36%. After transfer of >80 zygotes of each group into recipients, the respective percentages of full-term normal offspring resulting from frozen sperm or from freeze-dried sperm stored at +25, +4, and -196 degrees C were 36%, 0%, 7%, and 14%. These results demonstrate that the storage temperature significantly influenced the likelihood of term development of rats produced by injection of oocytes with freeze-dried spermatozoa. Chromosomal analysis of the rat spermatozoa in the ICSI oocytes indicated that chromosomal aberration in freeze-dried spermatozoa stored at +25 degrees C (100%) occurred more frequently than in frozen control spermatozoa (41%) and freeze-dried spermatozoa stored at -196 degrees C (35%), and the frequency of chromosomal aberrations in freeze-dried spermatozoa stored at +4 degrees C (65%) was the intermediate. In conclusion, rat spermatozoa freeze-dried and stored at +4 degrees C for 1 year are capable of participating in full-term development after ICSI.  相似文献   

15.
In order to work out a quarantine treatment for cactus pear fruit a factorial experimental plan was carried combining postharvest water dips at 20, 50, 54, 58 and 60 degrees C and storage at 1 degrees C for 3, 6 and 10 days. Cactus pear fruit cv 'Rossa' were artificially infested with Med. fly eggs (at least 20 eggs per fruit) then left in the lab at 25 degrees C for 4 days. Treatments took place by dipping the fruit at each water temperature for 2 mm. At each established time fruit was picked and checked for vital larvae and degree of chilling injury (CI). Probit 9 requirements were achieved in all cases when fruit was cold-stored for 10 days. When fruit was kept for 6 days the quarantine requirement was achieved only by dipping the fruit at 58 and 60 degrees C while none dip treatment was effective if fruit was stored for 3 days at 1 degrees C. All Fruit stored for 10 days at 1 degrees C showed severe CI symptoms and when kept for 6 days the same degree of CI was found on fruit dipped in water at 20, 58 and 60 degrees C. No CI was observed after 3 days at 1 degrees C. In conclusion only when fruit was dipped at 50-54 degrees C and stored for 10 days at 1 degrees C the probit 9 condition was attained with acceptable CI symptoms.  相似文献   

16.
AIMS: To determine survival and growth characteristics of Enterobacter sakazakii in infant rice cereal as affected by type of liquid used for reconstitution and storage temperature after reconstitution. METHODS AND RESULTS: A commercially manufactured dry infant rice cereal was reconstituted with water, apple juice, milk, or liquid infant formula, inoculated with a 10-strain mixture of E. sakazakii at populations of 0.27, 0.93, and 9.3 CFU ml(-1), and incubated at 4, 12, 21 or 30 degrees C for up to 72 h. Growth did not occur in cereal reconstituted with apple juice, regardless of storage temperature, or in cereal reconstituted with water, milk, or formula and stored at 4 degrees C. The lag time for growth in cereal reconstituted with water, milk, or formula was decreased as the incubation temperature (12, 21 and 30 degrees C) was increased. Upon reaching maximum populations of 7-8 log10 CFU ml(-1), in some instances populations decreased to nondetectable levels during subsequent storage which was concurrent with decreases in pH. CONCLUSIONS: Enterobacter sakazakii initially at very low populations can rapidly grow in infant rice cereal reconstituted with water, milk, or infant formula. SIGNIFICANCE AND IMPACT OF THE STUDY: Reconstituted infant rice cereal can support luxuriant growth of E. sakazakii. Reconstituted cereal that is not immediately consumed should be discarded or stored at a temperature at which E. sakazakii and other food-borne pathogens cannot grow.  相似文献   

17.
AIM: The influence of sample handling on soluble urokinase plasminogen activator receptor (suPAR) concentrations in serum and EDTA plasma was studied in 16 healthy premenopausal women. METHOD: Blood was collected in dry tubes and tubes containing EDTA and kept at 4 degrees C or 20 degrees C for 1, 3, 8, 24 or 72 hours before processing into serum or EDTA plasma. In addition, serum and EDTA plasma were frozen and thawed 1-8 times. All suPAR measurements were performed by ELISA. RESULTS: No significant differences were found between serum or EDTA plasma suPAR concentrations when whole blood samples were kept for 1, 3, 8 or 24 hours. Significantly higher suPAR levels were found in samples kept for 72 hours at 20 degrees C compared to samples processed into serum or EDTA plasma after short-term storage for no more than 24 hours after collection. No significant differences were observed when whole blood was kept at 4 degrees C for up to 72 hours. Repeated freezing and thawing had no significant effect on the serum and EDTA plasma suPAR levels. CONCLUSION: suPAR values in blood samples are dependent on the handling procedures of the samples. All samples of whole blood must be processed into EDTA plasma or serum within 24 hours if kept at 20 degrees C and within 72 hours if kept at 4 degrees C. However, repeated freezing/thawing cycles had no influence on suPAR values in the samples.  相似文献   

18.
Post-mortem spermatozoa recovery is an important technique for obtaining germplasm reserves from genetically valuable animals or endangered species. However, there are many factors that influence the outcome of this technique. We have studied the effect of the interval between animal's death and sperm recovery (0, 24 or 48 h) on the quality and freezability of ram spermatozoa from cauda epididymidis. Storage temperature of epididymis (room temperature or 5 degrees C) was also analysed. Spermatozoa were diluted with Tes-Tris-Fructose solution supplemented with egg yolk (10%) and glycerol (4%), and frozen using a programmable biofreezer (-20 degrees C/min). Pre-freeze and post-thaw sperm samples showed viable spermatozoa up to 48 h after the animal's death, although their quality declined significantly as post-mortem storage time increased. Epididymis sperm stored at 5 degrees C showed better motility and a lower percentage of abnormal forms than epididymis stored at room temperature after 24 and 48 h. The fertilizing ability of cauda epididymis ram spermatozoa obtained at 0 and 24h after the animal's death is similar to that of ejaculated spermatozoa. Therefore, a good protocol for post-mortem semen collection in rams when epididymal spermatozoa cannot be collected immediately, is to preserve the epididymis at 5 degrees C and process the samples in the first 24h after the animal's death.  相似文献   

19.
Apolipoprotein(a), apo(a), the specific multikringle glycoprotein constituent of lipoprotein(a), Lp(a), occurs in the plasma mostly bound to apoB100-containing lipoproteins but also in a free form. Often the properties of these products are determined after storage in the cold; yet limited information is available on their stability at low temperatures. To shed light on this subject, we examined the effect of two parameters, freezing and lyophilization, in either the absence or the presence of cryopreservatives. Lp(a)s each having a single apo(a) size isoform containing either 14 or 17 kringle (K) IVs were isolated from the plasma of healthy donors by combining density gradient ultracentrifugation and lysine-Sepharose column chromatography using solutions containing both antioxidants and proteolytic inhibitors. Apo(a) was obtained from parent Lp(a) by a mild limited reductive procedure. Either freezing at -20 degrees C or lyophilization in the presence of 5% sucrose did not change the electrophoretic, immunochemical, and lysine-binding properties of Lp(a) including its ability to generate free apo(a). Irrespective of source, apo(a) remained stable when either frozen at -20 and -80 degrees C or lyophilized in the presence of 125 mM trehalose. In all cases, the absence of cryopreservatives caused the samples to aggregate irreversibly. Thawed or reconstituted samples of both free and bound apo(a) kept at 4 degrees C under sterile conditions in the presence of antioxidants, proteolytic inhibitors, and cryopreservative exhibited no significant changes in properties within the time of observation. Both apo(a) isoforms gave comparable results. We conclude that apo(a), either free or bound, can be kept stable at low temperatures in the presence of appropriate cryopreservatives.  相似文献   

20.
The goal of modern transfusion therapy is to provide appropriate replacement therapy with blood components as opposed to whole blood for patients with specific hematologic deficiencies. A prerequisite of component therapy is, therefore, correct identification of the deficiency. Appropriate use of components avoids many of the hazards associated with the use of whole blood, and at the same time makes maximal use of this valuable resource. Blood components separated from whole blood soon after collection and appropriately stored can, in combination, provide all the factors present in fresh whole blood. Red cell concentrates prepared from multiple packs have a hematocrit of approximately 70%. They may be stored for up to 3 weeks at 4 degrees C and are recommended for most situations requiring red cell transfusions. Platelet concentrates, which can be stored for up to 72 hours at 22 degrees C, may be used for thrombocytopenic patients. Fresh frozen plasma, stored plasma, cryoprecipitated factor VIII, factor VIII concentrate and factor IX complex concentrate are available for the proper treatment of patients with hemorrhagic disorders due to coagulation factor deficiencies. Similarly, albumin and immune serum globulin are available for their oncotic and antibody properties respectively. Thus, the availability and appropriate use of the various blood products allows not only optimal transfusion therapy for each patient, but also fuller utilization of national blood resources.  相似文献   

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