Monoclonal antibody productivity and the metabolic pattern of perfusion cultures under varying oxygen tensions

The metabolic pattern and cell culture kinetics of high-cell-density perfusion cultures were compared under two different oxygen transfer conditions: oxygen limiting and not limiting. When oxygen was a limiting factor during perfusion culture, both specific glucose uptake and lactate production rates increased, compared to non-oxygen-limited condition, by about 60% and 30%, respectively. The specific glutamine uptake rate under oxygen-limited conditions was almost 4.0 times higher than that under non-oxygen-limited conditions. The activity of lactate dehydrogenase (LDH) released into the medium by the dead cells can be used as an indicator for the metabolic and physiological conditions related to oxygen limitation. There was a 3.2 times higher specific rate of LDH activity released by dead cells in oxygen-limited cultures than those in non-oxygen-limited cultures. The specific production rate of monoclonal antibody was not significantly affected by the oxygen transfer conditions during the rapid cell growth period, but it rapidly increased toward the end of perfusion cultures. The higher perfusion rate may have limited further cell growth during high-cell-density perfusion culture, because cell damage was caused by the hydrodynamic shear within a hollow fiber microfiltration cartridge installed to withdraw the spent medium and the waste metabolites. (c) 1993 John Wiley & Sons, Inc.     

A comparison of MS2-based label-free quantitative proteomic techniques with regards to accuracy and precision

The advent of algorithms for fragmentation spectrum-based label-free quantitative proteomics has enabled straightforward quantification of shotgun proteomic experiments. Despite the popularity of these approaches, few studies have been performed to assess their performance. We have therefore profiled the precision and the accuracy of three distinct relative label-free methods on both the protein and the proteome level. We derived our test data from two well-characterized publicly available quantitative data sets.     

Assessing constructed wetland functional success using diel changes in dissolved oxygen, pH, and temperature in submerged, emergent, and open-water habitats in the Beaver Creek Wetlands Complex, Kentucky (USA)

We assessed the functional success of restored wetlands by determining if the patterns in dissolved oxygen (DO), temperature, and pH were similar to those conditions observed in natural wetlands. The Beaver Creek Wetlands Complex consists of dozens of marshes and ponds built in a former Licking River floodplain, in the hills of east Kentucky, USA. In natural wetland ecosystems, aquatic primary production is highest in emergent and submerged vegetations zones; where daybreak dissolved oxygen (DO) is often near zero, and DO may rise to well over 100% saturation past mid-day. Open-water areas, dominated by phytoplankton, have less dramatic diel DO fluctuations—often without pre-dawn anoxia. Compared to open water, temperatures fluctuate less dramatically in vascular vegetation, due to shading and suppression of wind and waves. Measurements of ecosystem metabolism (diel changes in DO and pH) in three aquatic habitats of the constructed wetlands (emergent vegetation, submerged vegetation, open water) were compared to these natural ideals. In Beaver Creek Wetlands, water temperature patterns were not as dramatic as in natural habitats, nor did they did follow a similar trend. Waters in emergent vegetation (29.5 °C) were warmest; submerged vegetation coolest (26.5 °C); open-water intermediate (27.4 °C). Diel DO and pH patterns were not similar to natural habitats. Highest net primary production (NPP) and gross primary production (GPP) were measured in emergent vegetation waters (mean GPP = 7.58 g m⁻² d⁻¹); lowest in submerged vegetation (mean GPP = 5.48 g m⁻² d⁻¹); and intermediate in open-water (mean GPP = 6.95 g m⁻²d⁻¹). Diel pH changes were greatest in the highly productive emergent waters (median maximum daily difference of 0.36), and not as pronounced in submerged vegetation and open-water (median maximum change = 0.16 and 0.22, respectively). Water-column respiration was generally about double NPP. Like natural ecosystems, near anoxic DO concentrations were consistently measured in emergent and submerged plants before dawn; whereas open-water zones were generally >4 mg l⁻¹. These restored wetland systems may need more time to be functionally equivalent to natural marshes.     

Benthic community metabolism in four temperate stream systems: An inter-biome comparison and evaluation of the river continuum concept

Benthic community metabolism was studied on four stream systems located in different biomes in the United States: the eastern deciduous forest (Pennsylvania, PA, and Michigan, MI), the high desert (Idaho, ID), and the coniferous forest (Oregon, OR). Studies were designed to test the hypothesis advanced within the River Continuum Concept that a transition in community metabolism will occur from a predominance of heterotrophy in headwaters to a predominance of autotrophy in mid-sized reaches, with a return to heterotrophy further downstream. Both gross primary productivity (GPP) and community respiration (CR₂₄) increased with downstream direction on all systems. Net daily metabolism (NDM, or GPP – CR₂₄) shifted from heterotrophy (–NDM, GPP < CR₂₄) to autotrophy (+NDM, GPP > CR₂₄) with downstream direction at all sites, supporting the hypothesis. Annual metabolism in the most upstream reach of all sites was dominated by respiration; however, the farthest downstream reach was not necessarily the most autotrophic. Site-specific factors affected manifestation of the trend. Photosynthesis predominated annual metabolism in reaches (designated 1–4 in order of increasing size) 2–4 in ID, 3 and 4 in OR, and 4 in MI. In PA annual photosynthesis was slightly greater than respiration only at Station 3. Photosynthesis was predominant most consistently in ID and respiration most often in PA. About half the reaches that were heterotrophic annually were autotrophic at one or more seasons. Annual means of benthic GPP, CR₂₄ and NDM ranged from 0.16 to 3.37, 0.36 to 2.88 and –0.73 to 0.50 g O₂ · m² · d¹, respectively. Metabolic rates were usually high in PA and MI (and sometimes ID) and almost always lowest in OR. Parameters accounting for most variance in multiple linear regression analyses of the combined metabolism data from all sites were indicators of stream size, photosynthetically active radiation, temperature, and chlorophyll a concentration.     

A simple method to improve the precision and accuracy of gas-phase oxygen electrodes.

The precision and accuracy of the measurements using an oxygen gas-phase electrode thermostabilized in an ice bath were evaluated. The precision found was comparable with that obtained with the paramagnetic method and with Scholander analysis. The accuracy was comparable with that of the paramagnetic method.     

人工生境条件下几种红树植物的净初级生产力比较研究

选择3种红树植物海桑(Sonneratia caseolaris)、秋茄(Kandelia candel)和桐花树(Aegiceras corniculatum),每种分别按45%、30%和15%的面积比例种植于滩涂海水养殖塘.种植后连续2年对红树植物进行生态监测.结果表明,海桑增高457.0cm,基径增加86.1mm,成活率92.9%;桐花树高、基径分别增长2.1cm和36.5mm,成活率93.9%;秋茄成活率44.7%,增高20.4cm、基径增加2.4mm,说明在不受自然潮汐影响的人工生境条件下,海桑和桐花树对环境的适应能力强,生长较好,秋茄的生长适应性较差.据不同时期树高、基径与干、枝、叶、根的生物量,求得植物各器官生物量与树高、基径的回归方程,分析了3种红树植物的生物量与净初级生产力.海桑单位面积生物量5597.8g·m^-2,桐花树962.5g·m^-2,秋茄66.0g·m^-2.生物量在植物各器官的分配按大小排序,海桑为树干＞树枝＞树根＞树叶;桐花树为树叶＞树枝＞树干＞树根;秋茄为树干＞树根＞树叶＞树枝.单位面积净初级生产力海桑为7051.5g·m^-2,桐花树1105.8g·m^-2,秋茄93.0g·m^-2.高生产力伴随高归还量,凋落物归还量占净初级生产力的比重为海桑20.5%、桐花树15.4%、秋茄7.%.     

A comparison of methods for measuring primary productivity and community respiration in streams

Carbon dioxide and oxygen exchange procedures for measuring community metabolism (two open stream methods and three chamber methods) were compared on the same reach of a third-order stream. Open stream methods were complicated by high diffusion rates and yielded net community primary productivity estimates lower than those obtained with chamber methods. Chamber methods yielded variable productivity and respiration data. However, when normalized for chlorophyll a, productivity estimates from the chamber methods were within an expected range for the system. Balances of photosynthesis and respiration from the chamber methods were similar between methods and indicated that autotrophic or heterotrophic processes could dominate the system. Considerations in applying the various procedures are discussed.     

几种高通量定向筛选技术及其比较

在定向进化过程中,多样性的突变文库一旦构建好后,实验成败与否的关键就在于建立针对目的改造特征的高通量筛选方案。从突变文库中筛选需要的、功能进化的目的基因,关键在于是否有一种或几种高通量筛选技术。介绍了几种目前常用高通量定向筛选技术的基本原理,并对其特性进行了比较。     

Ceriometry,a combined method for chemical oxygen demand and dissolved oxygen (with a discussion on the precision of the winkler technique)

A ceriometric method is described in which Ce³⁺ salts are used for the determination of dissolved oxygen and Ce⁴⁺ salts for the determination of the chemical oxygen demand. The interference of COD in the O₂ determination, a common feature of most Winkler determinations, is corrected. The standard deviation is typically about 1% of the mean, and bias (inaccuracy) is very small.The method is simple, quick and reliable.The precision and accuracy of the Winkler method is discussed and compared with that of the Cerium method.     

Simultaneous determination of the accuracy and precision of closed-circuit cardiac output rebreathing techniques.

Foreign and soluble gas rebreathing methods are attractive for determining cardiac output (Q(c)) because they incur less risk than traditional invasive methods such as direct Fick and thermodilution. We compared simultaneously obtained Q(c) measurements during rest and exercise to assess the accuracy and precision of several rebreathing methods. Q(c) measurements were obtained during rest (supine and standing) and stationary cycling (submaximal and maximal) in 13 men and 1 woman (age: 24 +/- 7 yr; height: 178 +/- 5 cm; weight: 78 +/- 13 kg; Vo(2max): 45.1 +/- 9.4 ml.kg(-1).min(-1); mean +/- SD) using one-N(2)O, four-C(2)H(2), one-CO(2) (single-step) rebreathing technique, and two criterion methods (direct Fick and thermodilution). CO(2) rebreathing overestimated Q(c) compared with the criterion methods (supine: 8.1 +/- 2.0 vs. 6.4 +/- 1.6 and 7.2 +/- 1.2 l/min, respectively; maximal exercise: 27.0 +/- 6.0 vs. 24.0 +/- 3.9 and 23.3 +/- 3.8 l/min). C(2)H(2) and N(2)O rebreathing techniques tended to underestimate Q(c) (range: 6.6-7.3 l/min for supine rest; range: 16.0-19.1 l/min for maximal exercise). Bartlett's test indicated variance heterogeneity among the methods (P < 0.05), where CO(2) rebreathing consistently demonstrated larger variance. At rest, most means from the noninvasive techniques were +/-10% of direct Fick and thermodilution. During exercise, all methods fell outside the +/-10% range, except for CO(2) rebreathing. Thus the CO(2) rebreathing method was accurate over a wider range (rest through maximal exercise), but was less precise. We conclude that foreign gas rebreathing can provide reasonable Q(c) estimates with fewer repeat trials during resting conditions. During exercise, these methods remain precise but tend to underestimate Q(c). Single-step CO(2) rebreathing may be successfully employed over a wider range but with more measurements needed to overcome the larger variability.     

2000—2015年拉萨河流域NPP时空变化及驱动因子

拉萨河流域是西藏政治、经济和文化核心区,具有青藏高原典型的高寒湿地及西藏主要的耕作区,受人类活动及气候变化的影响,流域生态安全及生态健康面临威胁和破坏。植被净初级生产力（NPP）作为衡量生态系统健康的重要指标,能够反映生态系统的可持续发展。利用2000-2015年MODIS-NDVI数据,基于光能利用率模型估算了拉萨河流域的NPP,分析了NPP的时空格局、动态变化特征及NPP与气候因子的相关性,探讨了影响NPP变化的驱动因子并明确了驱动分区。结果表明：近16年拉萨河流域NPP年均值为165.614 gC m^-2 a^-1,总体分布具有明显的空间异质性,与该地区植被类型的分布规律相似,不同植被类型的NPP存在差异。NPP变化在总体上呈下降趋势,平均年变化趋势斜率为-1.804 gC m^-2 a^-1。NPP与气温和降水的相关性具有明显的地域性差异,草本湿地与气温呈显著负相关,灌丛与降水呈显著正相关。NPP变化受气候因子驱动的区域占比20.81%,非气候因子占比79.19%。     

Response of river metabolism to restoration of flow in the Kissimmee River, Florida, U.S.A.

1. The single station diel oxygen curve method was used to determine the response of system metabolism to backfilling of a flood control canal and restoration of flow through the historic river channel of the Kissimmee River, a sub‐tropical, low gradient, blackwater river in central Florida, U.S.A. Gross primary productivity (GPP), community respiration (CR), the ratio of GPP/CR (P/R) and net daily metabolism (NDM) were estimated before and after canal backfilling and restoration of continuous flow through the river channel. 2. Restoration of flow through the river channel significantly increased reaeration rates and mean dissolved oxygen (DO) concentrations from <2 mg L⁻¹ before restoration of flow to 4.70 mg L⁻¹ after flow was restored. 3. Annual GPP and CR rates were 0.43 g O₂ m⁻² day⁻¹ and 1.61 g O₂ m⁻² day⁻¹ respectively, before restoration of flow. After restoration of flow, annual GPP and CR rates increased to 3.95 O₂ m⁻² day⁻¹ and 9.44 g O₂ m⁻² day⁻¹ respectively. 4. The ratio of P/R (mean of monthly values) increased from 0.29 during the prerestoration period to 0.51 after flow was restored, indicating an increase in autotrophic processes in the restored river channel. NDM values became more negative after flow was restored. 5. After flow was restored, metabolism parameters were generally similar to those reported for other blackwater river systems in the southeast U.S.A. Postrestoration DO concentrations met target values derived from free flowing, minimally impacted reference streams.     

Effect of inorganic sediment on whole-stream productivity

Sediment from agricultural, logging, and mining activities impairs more miles of rivers and streams in the United States than any other type of pollutant, including bacteria, nutrients, oxygen-depleting substances, and metals. However, specific impacts of sediment to streams have not been well studied or understood. To study the effect of inorganic sediment on plant and animal communities in stream ecosystems, we added clay to outdoor experimental streams 520 m long and 3.5 m wide at the Monticello Ecological Research Station (MERS). The streams take water from the Mississippi River and are designed to represent higher order streams in the upper midwestern United States. The sediment loading rates were 300, 200, 100, and 50 mg l^–1. Our first dosing period (mid-August to November 1994) began at the start of a fall bloom in autotroph productivity, and the second (May to August 1995) began before the summer communities were established. During both treatment seasons, the addition of clay significantly increased turbidity and sedimentation, and decreased light penetration in treated streams corresponding roughly to 30–35, 25–30, 15–20, and 5–10 NTU, respectively. In general, the macrophyte and periphyton communities responded quickly after only a few weeks exposure to the sediment additions. Whole-stream respiration was significantly lower in treated streams, decreasing as the amount of sediment added increased. Periphyton biomass (chl a) on tiles and percent cover of macrophytes was significantly lower in treatment streams than in controls. In contrast to expectations and previous findings in two sets of field streams, total whole-stream productivity in the MERS streams was not significantly lower in streams receiving sediment loads than in control streams because the overall photosynthetic efficiency by the plant community compensated for the loss in irradiance.     

Hypoxic fed-batch cultivation of Pichia pastoris increases specific and volumetric productivity of recombinant proteins

High cell density cultivation of Pichia pastoris has to cope with several technical limitations, most importantly the transfer of oxygen. By applying hypoxic conditions to chemostat cultivations of P. pastoris expressing an antibody Fab fragment under the GAP promoter, a 2.5-fold increase of the specific productivity q(P) at low oxygen supply was observed. At the same time the biomass decreased and ethanol was produced, indicating a shift from oxidative to oxidofermentative conditions. Based on these results we designed a feedback control for enhanced productivity in fed batch processes, where the concentration of ethanol in the culture was kept constant at approximately 1.0% (vv(-1)) by a regulated addition of feed medium. This strategy was tested successfully with three different protein producing strains, leading to a three- to sixfold increase of the q(P) and threefold reduced fed batch times. Taken together the volumetric productivity Q(P) increased 2.3-fold.     

A comparison of staining techniques to improve precision of age estimation from fish otoliths

A total of 30 pairs of sagitta otoliths of horse mackerel Trachurus trachurus were sampled from three length-classes, and five staining methods were applied to each pair. Ten age estimates were obtained per fish, method and length-class. Significant differences in age estimates were found between methods, due to the variable enhancement in the otolith pattern produced by each method. A method using amido-schwartz as the staining reagent produced the best results in terms of precision of the age estimates.     

Labor productivity: A biophysical definition and assessment

A model of energy analysis is presented to study the concept of labor productivity from a biophysical perspective. It is argued that current methods of defining and assessing labor productivity in the fields of work physiology and input/output energy analysis are relatively poor operational tools for assessing productivity in the economy and society. We propose to adopt society as the hierarchical level of analysis rather than the individual, as labor productivity can best be studied as a function of parameters related to the technological development of society. Parameters considered are: the ratio exosomatic/endosomatic energy used in society, the ratio working/non-working population, the return on the circulating energy investment, and the profile of human time allocated to the economic process. The links between patterns of human time allocation, population structure, standard of living, technological development, and demand on natural resources are analyzed. The results suggest that the role and meaning of human labor differ widely in societies with different levels of technological development.     

A new method to measure the haemoglobin oxygen saturation by the oxygen electrode

We describe a new polarographic method to measure the haemoglobin oxygen saturation in whole blood, employing up to 10 μl of sample in a standard case. The measurement is done in an anaerobic staineless-steel cuvette (1 ml) recording three oxygen tension values: (1) that of an air-equilibrated buffer before the addition of the sample; (ii) that after the addition of the sample; and (iii) that after the addition of an oxidant. The haemoglobin oxygen saturation is then calculated from the three oxygen tension values, the volume of the reagents, and solubility coefficient of oxygen. This method is simple, inexpensive and accurate, and correlates well with other standard methods.