CELL AND GROWTH CHARACTERISTICS OF TYPES A AND B OF EMILIANIA HUXLEYI (PRYMNESIOPHYCEAE) AS DETERMINED BY FLOW CYTOMETRY AND CHEMICAL ANALYSES1

Two morphotypes of Emiliania huxleyi (Lohmann 1902) Hay et al. 1967, types A and B, known to be unequally distributed in the oceans, were grown in dilution cultures at a range of photon flux densities (PFDs) (1.5–155 μmol photons·m^?2·s^?1) and two temperatures (10° and 15° C). Calcite carbon and organic carbon content of the cells as well as instantaneous growth rate, cell size, chlorophyll fluorescence, and light-scatter properties clearly depended on growth conditions and differed considerably for the two morphotypes. The ratio between calcite carbon and organic carbon production showed an optimum of 0.65 in E. huxleyi type A cells at PFD = 17.5. The ratio increased slightly with a temperature increase from 10° to 15°C but remained < 1.0 at both temperatures in light-limited cells. In contrast, calcite carbon production exceeded organic carbon production (ratio: 1.4–2.2) in phosphate-deprived cultures. Emiliania huxleyi type B generally showed a higher calcite carbon/organic carbon ratio than E. huxleyi type A, but the relation with PFD was similar. The content of calcite carbon and organic carbon as well as the instantaneous growth rate, cell size, chlorophyll fluorescence, and light-scatter properties showed large diel variations that were closely related to the division cycle. Our results show the importance of mapping the structure of any sampled cell population with respect to the phase in the cell division cycle, as this largely determines the outcome of not only “per cell” measurements but also short time (less than 24 h) flux measurements. For instance, dark production of calcite by E. huxleyi was negatively affected by cell division. Slowly growing (phosphate-stressed) cultures produced calcite in the light and in the dark. In contrast, rapidly growing cultures at 10°C produced calcite only in the light, whereas in the dark there was a significant loss of calcite due to dissolution.     

A COMPARISON OF NUCLEAR DRY WEIGHTS DETERMINED BY CHEMICAL AND BY INTERFEROMETRIC METHODS

1. The nuclei of cells from the thymus of the calf were isolated by three different techniques; the citric acid, the sucrose-calcium chloride, and the non-aqueous. 2. The mean dry weights of the nuclei were determined by chemical methods and by microscopic interferometry. There was a close correlation between the results from the interferometric and chemical methods. 3. The range of values about that mean was determined in each sample: the nuclei isolated in aqueous media contained approximately 45 per cent less material than those isolated in non-aqueous media. 4. The variations in dry weight with varying nuclear type are discussed. 5. The possible relationship between DNA content and dry weight is discussed.     

CELL-PRODUCTION RATES ESTIMATED BY THE USE OF VINCRISTINE SULPHATE AND FLOW CYTOMETRY

A new method for the evaluation of cell production rates combining flow cytometry (FCM) and the stathmokinetic method using vineristine sulphate (VS) has been used for the analysis of three aneuploid ascites tumours at different stages of growth. Using this technique it was possible to estimate the well-known decrease in cell production rates of ageing ascites tumours. The percentage of normal host cells in the aneuploid tumours studied was easily determined by FCM prior to the calculation of the tumour cell-production rates. A correlation was found between the percentage of tumour cells in the S phase and the tumour cell-production rate. This correlation is probably explained by the gradual transfer of proliferating cells in S phase to resting G₁ and G₂ phases with increasing tumour age.     

CHARACTERIZATION OF OCEANIC PHOTOSYNTHETIC PICOEUKARYOTES BY FLOW CYTOMETRY1

To interpret flow cytometric data that are routinely obtained on natural oceanic communities, 23 strains of photosynthetic picoeukaryotes belonging to four classes (Prasinophyceae, Chlorophyceae, Pelagophyceae, and Prymnesiophyceae) and six pigment types were investigated for their light scattering in the forward and right-angle directions, chlorophyll fluorescence, and DNA content as measured by flow cytometry. Cell she was assessed by Coulter counter, and pigment composition was measured by reverse-phase high-performance liquid chromatography. The size and GC% of the nuclear genome of cultured picoeukaryotes was measured from the fluorescence of DNA-specific dyes. Using these two parameters, we could discriminate species within pigment groups. DNA staining of preserved natural samples may also prove useful in discriminating cooccurring populations in situ as long as the communities are not too complex. Using the relationships that we established between size and light-scattering properties of the cells, we estimated equivalent diameters of picoeukaryotes in natural populations to be between 1.3 and 2 μm. Chlorophyll a content was between 6 and 16 fg·cel^?1 as calculated from relationships that we established between chlorophyll a content and red fluorescence of the cultured strains. With respect to size, chlorophyll a content, and pigment composition, Pelagomonas sp. strains (Pelagophyceae) appeared to be the most representative of the natural communities in subtropical ocean waters. In contrast, green coccoid strains, which often outcompete other strains in culture, might only be minor contributors to these communities.     

小儿急性白血病细胞DNA、蛋白质含量及其临床意义

用流式细胞术（ＦＣＭ）测定了４５例不同病期急性白血病患儿和１３例非白血病患儿骨髓细胞ＤＮＡ及蛋白质含量,结果显示：１．急性白血病患儿的ＤＮＡ非整倍体检出率为４１．２％,其中ＡＬＬ为３３．３％,ＡＮＬＬ为６０％;２．ＡＬＬ组和ＡＮＬＬ组的ＤＮＡ合成期细胞百分数（Ｓ％）显著低于非白血病组,而ＣＲ组与非白血病组之间的Ｓ％差异无显著性;３．ＡＮＬＬ组的ＰｒＩ显著高于ＡＬＬ组、非白血病组及ＣＲ组,ＡＬＬ的ＰｒＩ显著低于ＣＲ组和非白血病组,而非白血病组与ＣＲ组的ＰｒＩ差异无显著性。四组之间ＬＰＣ－Ｆ差异无显著性;４．在对１１例ＣＲ病人进行的连续观察过程中,５例病人检出ＤＮＡ非整信体,其中４例于检出ＤＮＡ非整倍体后３３～７４天复发。     

流式细胞光度术DNA分析在非何杰金氏淋巴瘤预后中的意义

用流式细胞光度术对一组非何杰金氏淋巴瘤(NHL)存档石蜡标本进行DNA分析。DNA异倍体肿瘤占总例数的47.4％,高、中度恶性组异倍体肿瘤的比例(76.8％和51.9％)分别高于低度恶性组(17.6％)(P<0.05)。S期细胞比例(synthetic phase fraction,SPF)随组织学恶性程度升高而增大,低、中和高度恶性淋巴瘤平均SPF值分别是6.5(0.6—18.8)％,13.5(3.2—37.3)％和23.4(4.4—41.4)％。二倍体和异倍体肿瘤患者的生存率无差别。全部病例中高SPF值肿瘤患者具有生存短的趋向(P=0.1)。低度恶性组内也存在此种趋向(P=0.08)。上述结果表明流式细胞光度术DNA分析对估价非何杰金氏淋巴瘤生物学行为是一种不依赖形态学的有用方法。     

DETERMINATION OF NON-PROLIFERATING CELLS IN CULTURE BY COMBINING FLOW CYTOMETRY WITH STATHMOKINETICS

Colcemid was added to the growth medium of L-cells in monolayer culture. The proliferating cells continued their progression through the cycle up to metaphase, where they were arrested. At different times after Colcemid addition the cells were trypsinized. suspended immediately in a solution of the DNA-specific fluorescent dye Hoechst 33258 and analysed with a flow cytometer. The histograms were evaluated to give the fraction of cells in the 2c peak as a function of time after Colcemid addition. The flux into the 2c compartment being interrupted, the peak content decreased until all proliferating (G₁) cells had entered S-phase. With increasing cell density or with increasing time after serum deprivation an increasing fraction of cells remained in the 2c peak at times greater than the normal G₁ duration. The possibility of applying this method to the determination of non-proliferating cells in a population is discussed.     

CD11b/DNA双参数流式细胞术检测HL-60细胞分化的细胞周期

目的采用双参数流式细胞术研究全反式维甲酸(alltransretinoidacid,ATRA)诱导人类急性早幼粒白血病细胞HL-60细胞分化的细胞周期。方法HL-60细胞经分化诱导剂ATRA(终浓度为1μmol/L)诱导不同时间点后,利用CD11b/DNA双参数流式细胞术同时检测分化细胞表面抗原CD11b的表达及分化细胞DNA含量。结果HL-60细胞经ATRA诱导后,细胞表面分化抗原CD11b表达明显升高,细胞阻滞于G0/G1期,且CD11b阳性细胞主要位于G0/G1期。结论CD11b/DNA双参数流式细胞术能简便,快速,直观地检测细胞分化的细胞周期。     

RAPID DETECTION of STAPHYLOCOCCUS AUREUS IN FOOD BY FLOW CYTOMETRY

This report describes the detection of Staphylococcus aureus in buffer and in several kinds of food by flow cytometry. Fluorescein isothiocyanate conjugated anti-protein A antibodies were used in a 4-h procedure to label cells, 10⁵-10⁶ cells/mL are needed. the use of single parameter, green fluorescence, enabled specific differentiation of S. aureus from other bacteria including 11 Staphylococcus species. the flow cytometric method can detect S. aureus in food samples after 48-h enrichment in trypticase soy broth with 10% NaCl. As low as 2 S. aureus cells present in 10 mL enrichment broth could grow to a population density detectable by the flow cytometric method after enrichment. This method was faster and less laborious than the conventional BAM (Bacteriological Analytical Manual) or AOAC (Association of Official Analytical Chemists) methods, and could be automated for analysis of S. aureus in food.     

应用流式细胞仪定量评估人—鼠杂交瘤株细胞抗体生成功能稳定性的研究

用FACS分析和Luria-Delbrǔck方程计算淋巴细胞杂交瘤株细胞丧失抗体生成功能的突变机率(μ),以评估杂交瘤的稳定性。结果表明μ值大,则杂交瘤不稳定,易丧失抗体生成功能,反之则杂交瘤比较稳定。实验证明所建立的方法能反映杂交瘤稳定性的程度,具有实验少人为主观因素干扰,统计意义强,适用范围广等优点。     

A COMPARISON OF PERMEABILITY IN PLANT AND ANIMAL CELLS

Quantitative studies show a striking agreement between frog skin and plant tissues in respect to certain important aspects of permeability, antagonism, injury, recovery, and death.     

FOUR-STRANDED DNA AS DETERMINED BY ELECTRON MICROSCOPY

Pneumococcus DNA, of weight-average molecular weight 1.6 million by light scattering, had a weight-average length of 4300 A by electron microscopy. Thus, the average mass per unit length was 370 molecular-weight units per A, or approximately two times that expected (208) for a Watson-Crick double helix. This corresponds to an average of 3.6 strands per molecule, which is close to that obtained by other methods. Morphologically, all the particles in the micrographs were relatively stiff, and had a cross-sectional height of 20 to 30 A. Some divided into two stiff branches of the same height, apparently double helical. Where the branches combined into one (minimally four-stranded) structure they apparently lay side by side in close association.     

大蒜油抗肿瘤作用的流式细胞光度术分析

用流式细胞光度术（ｆｌｏｗｅｙｔｏｍｅｔｒｙ,ＦＣＭ）+Ｓ０．２ｍｌ／日。吐温对照组ｌＰ吐温８００．２ｍｌ／日,以排除大蒜油制剂中溶剂（吐温８０）的影响。实验组ｌＰＧＯ１００ｍｇ／ｋｇ／．日。于连续４次注射后的６ｈ,２、３、５、１０天及１０次注射ＧＯ后７天取材,行ＦＣＭ样品制备。三、ＦＣＭ样品制备与测定中国中医研究院基础所腹水癌细胞用ＰＢＳ洗涤,７０％酒精固定,ＲＮａｓｅ消化细胞中的ＲＮＡ,ＰＩ染色。用腹腔内淋巴细胞作标准二倍体细胞。用４２０型荧光激活细胞分选仪（美国Ｂｅｃｔｏ－Ｄｉｃｋｓｏｎ公司产品）测单个肿瘤细胞的ＤＮＡ含量,用组方图显示肿瘤细胞ＤＮＡ倍体性质。图中第一个是ＤＮＡ二倍体（ＺＣ）峰,处于该峰的细胞为Ｇ;／Ｇ。期细胞,第二个峰为４ＣＤＮＡ峰,该处细胞为Ｇ。－Ｉ－－Ｍ期细胞,从２。到４。之间的细胞为Ｓ期细胞。大于４。ＤＮＡ峰细胞为高倍体细胞‘’：。根据不同峰值大小判定用药后肿瘤细胞倍体性质的变化。结果ＮＳ与吐温对照组肿瘤细胞都为非整倍体性,且呈多倍体性（图１,２）。连续４次给药后６ｈ,绝大部分多倍体肿瘤细胞被杀伤,残存肿瘤细胞以二倍体为主,多倍体肿瘤细胞峰值显著下降,甚至在组方图上几乎显不出多倍     

流式双参数检测在肿瘤增殖,分化与凋亡中的应用

流式细胞术已在生物医学研究中有广泛应用,双参数流式术显著特点是给同一细胞群提供双重信息。本文成功地将这一先进技术用于食管癌增殖、分化与凋亡的检测,显示流式双参数的广阔应用前景。     

0#柴油对斑马鱼基因组DNA和环境DNA遗传毒性的RAPD比较

研究利用随机扩增多态性DNA (Random Amplified Polymorphic DNA, RAPD)技术, 以斑马鱼基因组DNA和其养殖水体中的环境DNA (environmental DNA, eDNA)为模板, 检测0#柴油可溶性组分对斑马鱼(Danio rerio)遗传毒性的影响。结果显示, 通过基因组DNA和eDNA扩增的RAPD图谱均可检测到0#柴油对斑马鱼的遗传毒性。在未受到柴油暴露时, 斑马鱼基因组DNA和水环境中eDNA在96h内的RAPD图谱均无明显变化; 在不同浓度的柴油暴露下, 随着暴露时间(0、24h、48h、72h、96h)延长, 基因组DNA和eDNA的多态性位点减少, 模板稳定性降低; 随着柴油浓度(15%、50%、100%)的增加, 基因组DNA和eDNA的多态性位点也减少, 模板稳定性降低。这表明0#柴油对斑马鱼基因组DNA和eDNA的遗传毒性均呈现时间-效应和浓度-效应关系, 并且无论以斑马鱼基因组DNA还是eDNA为模板, 柴油暴露组和未进行暴露的对照组的RAPD扩增图谱条带变化趋势一致。研究结果为通过RAPD技术检测柴油对水生生物的遗传毒性提供了新的研究思路和技术手段。     

PHYLOGENETIC COMPARISON OF LARGE NUCLEAR DNA CONTENTS OF DIFFERENTIATED CELLS IN THE ROOTS OF EQUISETUM,TRADESCANTIA, AND HORDEUM

Nuclear DNA of meristematic, epidermal and root cap cells from the roots of three vascular plants—the cryptogam, Equisetum hyemale L, and the phanerogams, Tradescantia Clone 02 and Hordeum vulgare L.—was measured with quantitative Feulgen microspectrophotometry. Epidermal cells of all three species and root cap cells in both phanerogams contained up to 8fold the amount of nuclear DNA found in their respective meristematic telophase nuclei. In general, the large amounts of nuclear DNA parallel development and differentiation in the epidermis regardless of phylogeny, habitat, or degree of domestication. However, comparisons of the increase in nuclear DNA contents in the various epidermal cell types among these three species suggest that the mechanisms giving rise to these increases may differ phylogenetically and may represent another character in which cryptogams and phanerogams diverged in their evolution.