作者承认国家科学基金会研究与创新新兴前沿（第1332341号），用于资助这项研究。作者还要感谢Andrew Grieshop博士以及LabJack和DAQFactory在线支持社区在整个过程中提供的帮助和帮助。

<ol>
	<li>Liu, X., Clarens, A. F., Colosi, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Algae+biodiesel+has+potential+despite+inconclusive+results+to+date.">Algae biodiesel has potential despite inconclusive results to date.</a> Bioresour. Technol. 104, 803-806 (2012).</li><li>Chisti, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biodiesel+from+microalgae.">Biodiesel from microalgae.</a> Biotechnol. Adv. 25 (3), 294-306 (2007).</li><li>Lardon, L., H&#233;lias, A., Sialve, B., Steyer, J. -. P., Bernard, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Life-cycle+assessment+of+biodiesel+production+from+microalgae.">Life-cycle assessment of biodiesel production from microalgae.</a> Environ. Sci. Technol. 43 (17), 6475-6481 (2009).</li><li>Henry, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experiments+on+the+Quantity+of+Gases+Absorbed+by+Water,+at+Different+Temperatures,+and+under+Different+Pressures.">Experiments on the Quantity of Gases Absorbed by Water, at Different Temperatures, and under Different Pressures.</a> Philos. Trans. R. Soc. London. 93 (1803), 29 (1803).</li><li>Wang, X., Hao, C., Zhang, F., Feng, C., Yang, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inhibition+of+the+growth+of+two+blue-green+algae+species+(Microsystis+aruginosa+and+Anabaena+spiroides)+by+acidification+treatments+using+carbon+dioxide.">Inhibition of the growth of two blue-green algae species (Microsystis aruginosa and Anabaena spiroides) by acidification treatments using carbon dioxide.</a> Bioresour. Technol. 102 (10), 5742-5748 (2011).</li><li>Juneja, A., Ceballos, R. M., Murthy, G. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+Environmental+Factors+and+Nutrient+Availability+on+the+Biochemical+Composition+of+Algae+for+Biofuels+Production:+A+Review.">Effects of Environmental Factors and Nutrient Availability on the Biochemical Composition of Algae for Biofuels Production: A Review.</a> Energies. 6 (9), 4607-4638 (2013).</li><li>Bernard, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hurdles+and+challenges+for+modelling+and+control+of+microalgae+for+CO+2+mitigation+and+biofuel+production.">Hurdles and challenges for modelling and control of microalgae for CO 2 mitigation and biofuel production.</a> J. Process Control. 21 (10), 1378-1389 (2011).</li><li>Guest, J. S., van Loosdrecht, M. C. M., Skerlos, S. J., Love, N. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lumped+Pathway+Metabolic+Model+of+Organic+Carbon+Accumulation+and+Mobilization+by+the+Alga+Chlamydomonas+reinhardtii.">Lumped Pathway Metabolic Model of Organic Carbon Accumulation and Mobilization by the Alga Chlamydomonas reinhardtii.</a> Environ. Sci. Technol. 47 (7), 3258-3267 (2013).</li><li>Packer, A., Li, Y., Andersen, T., Hu, Q., Kuang, Y., Sommerfeld, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+and+neutral+lipid+synthesis+in+green+microalgae:+A+mathematical+model.">Growth and neutral lipid synthesis in green microalgae: A mathematical model.</a> Bioresour. Technol. 102 (1), 111-117 (2011).</li><li>Oren, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+hundred+years+of+Dunaliella+research:+1905-2005.">A hundred years of Dunaliella research: 1905-2005.</a> Saline Systems. 1, 2 (2005).</li></ol>

作者没有什么可以披露的。

该PBR系统提供监测和控制实验室规模的藻类动力学生长实验的能力，允许来自用于量化生长的实验测定的更多可重复的结果。然而，了解传感器测量的局限性和不确定性对于确保传感器读数准确反映反应器条件至关重要。这种理解包括与传感器相关的测量原理的基本知识，校准的过程和频率，测量不确定度以及传感器可以测量甚至不能测量的内容。例如，这里描述的光传感器的电响应不是均匀地分布在可见光谱范围内，并且可能需要将某些校正因子应用于传感器输出，这取决于如何分析该传感器数据。 温度水平和变化也是非常重要的，因为温度的变化可以大大的变化注意传感器响应。了解可能影响传感器读数的潜在干扰也至关重要;这种干扰可能是来自建筑物的环境电气噪声，或者可能来自测量环境（ 例如，钠离子可以大大影响pH值超过10的pH读数）。此外，将多个探针浸入溶液中，特别是高度离子和导电的盐溶液也是潜在的干扰源。测量pH（或离子强度，溶解氧，溶解的CO 2 等 ）的电极对环境电气噪声特别敏感，并且可以容易地扰动。用于保护电极信号的信号调理不能保证其他因素不会干扰探头读数。作为质量控制的一部分，应使用其他实验室设备，如手持式pH探针，手持光谱仪和温度计，以验证t他的传感器读数，并确保系统设置和运行正常。 必须解决的另一个限制是藻类和/或培养环境对传感器的可能影响。例如，如果藻类碎片或泡沫覆盖光传感器的光电二极管接收器，读数将受到影响。类似地，pH电极是非常敏感的，需要特别小心以确保准确的读数。这些电极通过测量由于H +离子的积聚而在内部连接处的电压差而起作用;需要探针内的水合缓冲层以保持精确的测量12 。取决于反应器内的条件，该层将会脱落，并且当探头浸没时，传感器的响应可能会在实验过程中发生变化。在初步测试中，在20天的实验过程中，pH电压输出没有漂移超过约0.2个pH单位，但应进行进一步评估，以表征传感器响应的这种变化，并建立最大的实验运行时间，特别是如果需要精细的pH调节/定量。 许多现有的用于分析藻类生长的实验室规模的PBR系统不会像需要一样严密地监控和控制内部文化环境，以确定不同因素如何影响藻类生长，因为以这种方式建立系统可能具有挑战性。该协议可以通过提供用于构建具有实时监控的PBR的分步说明来帮助实现更多的受控实验。此外，这种实时数据不仅可以用于更好地控制实验条件，而且可以用于估计生长动力学（ 例如，光密度读数作为一般生长速率的参考）。 受控的实验系统可以帮助藻类研究更加可重现。台秤PBR监测和控制的水蛭可以通过最小化实验设计中的非预期人为因素来提高实验效率，并有助于推动使藻类生物燃料成为可持续的替代燃料来源。

由于对全球气候变化和有限化石燃料资源的担忧日益增加，各国政府一直在制定减少化石燃料消费政策并鼓励开发新的可持续运输燃料。美国环境保护局制定了可再生能源燃料标准（RFS），要求到2022年，每年1400亿加仑的美国运输燃料组合中有36个来自可再生燃料。创新和转型技术将是必要的，以满足这些和未来可再生能源标准1 。 使用微藻类生物燃料具有帮助满足国家RFS同时减少温室气体排放的潜力2 。与基于陆地粮食作物的第一代生物燃料相比，微藻类生物燃料具有几个优点，如玉米和大豆。与第一代生物燃料不同，藻类b生物燃料消耗较少的土地，水和与食物有关的资源，因为藻类可以全年耕种，在荒地上使用盐水或废水。微藻与陆地作物相比具有较高的生长速率，可以积聚高水平的脂质，可以容易地转化为生物柴油3 。目前，由于由藻类培养，脂质分离和脂质精炼成生物柴油的能源密集型生产过程的高成本，不存在工业规模的藻类 - 生物燃料工厂。需要更多的研究来使这些过程更有效率和可持续性。 用于在人造环境中生产光营养微生物的光学透明封闭装置的PBR被认为是最有希望的培养方法之一3 。然而，目前的设计仍然缺乏使藻类对生物燃料生产过程所需的体积生产力ess更有效率和经济吸引力4 。强大的数学模型，考虑光照辐射和衰减，营养物质和二氧化碳的运输，以及微藻的生长可以极大地促进PBR设计和操作的优化。为了确定这些优化模型的物种特异性生长参数，需要进行大规模生长实验。 动力学测试需要仔细监测和控制实验装置，以防止意外的生长抑制剂。鉴于藻类的光合特性（ 即它们的二氧化碳消耗和光的吸收），维持受控条件在实验室规模的PBR中特别困难。如等式1所示 ，生长培养基中溶解的CO 2的量通常表示为<img alt="方程" src="/files/ftp_upload/55545/55545eq1.jpg" /> （ 等式2 ）将至少为a功能:1）CO 2分压和亨利均衡常数，这决定了溶解在溶液中的气体量（ 等式3 ）; 2）生长培养基的初始化学成分，影响碳酸根离子的形态和活性，pH值（ 等式4和5 ）。和3）温度，影响等式3-5 5 。 <img alt="方程" src="/files/ftp_upload/55545/55545eq2.jpg" /> <img alt="方程" src="/files/ftp_upload/55545/55545eq3.jpg" /> <img alt="方程" src="/files/ftp_upload/55545/55545eq4.jpg" /> <img alt="方程" src="/files/ftp_upload/55545/55545eq5.jpg" /> <img alt="方程" src="/files/ftp_upload/55545/55545eq6.jpg" />碳的各个阶段和化学形态对测量和维持PBR中溶解碳浓度一致性构成挑战保持其他条件不变（ 例如，随着藻类消耗CO 2 ，pH增加，并且增加溶解的CO 2底物可能导致抑制生长的酸性环境）。 在藻类动力学测试中控制条件的另外一层复杂性涉及PBR内的光强度。 PBR内的平均光强度不仅是入射光强度，还包括设计（ 如材料，形状，深度和混合），藻类生物量成分（特别是叶绿素）的吸光度，藻细胞的散射性质。随着藻类生长，平均光强度将会降低。光强度的这种变化，无论是由总细胞和生物量的增加，每个细胞的叶绿素含量的增加还是两者引起的，都可能最终诱导代谢反应，如叶绿素产量的增加细胞每个细胞或使用碳水化合物和脂质储存产品的能量7 。连续监测反应堆内的光强，提供了宝贵的信息。该数据有助于确保条件保持在指定范围内，如果与其他测量（ 即生物量，叶绿素浓度，反应器深度，入射光等 ）结合使用，可用于帮助估计藻类生长和吸光度参数。 了解藻类在指定条件下如何生长需要在实验室规模的动力学实验中监测pH，溶解的CO 2 ，光强度和温度。许多藻类生长设置没有装备来监测校准动力学模型所需的条件，使建模过程非常具有挑战性8 。虽然许多公司提供具有自动化和控制功能的台式PBR，但这些台式机e设置可能非常昂贵（约$ 20,000），并且可能不适应给定研究问题的所有实验考虑。 建立批量实验的控制反馈系统的第一步是实时数据采集。本文旨在演示如何构建和设置配备连续光，pH和温度监测的工作台式PBR。这种实时监控设置可以帮助确保实验条件保持在所需范围内，由研究者自行决定。尽管该协议没有详细说明具体的控制机制，但是这些分步指令为更高级的控制反馈可以实现所需的数据采集框架提供了基础。

<table border="0" cellpadding="0" cellspacing="0" width="782">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Cast acrylic sheets</td>
			<td style="width:133px;">McMaster Carr</td>
			<td style="width:135px;">8560K244</td>
			<td style="width:345px;">7/8&#39;&#39; thick, 12x36&#39;&#39;, optically-clear, the size of sheets purchased will depend on reactor dimensions.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:169px;">Acrylic cement</td>
			<td style="width:133px;">McMaster Carr</td>
			<td style="width:135px;">7517A4</td>
			<td style="width:345px;">Scigrip plastic pipe cement, #4SC nonwhitening for acrylic. Not needed if gaskets and screws are used for PBR assembly.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Acrylic cement applicator needle</td>
			<td style="width:133px;">McMaster Carr</td>
			<td style="width:135px;">75165A136</td>
			<td style="width:345px;">Acrylic cement applicator needle, 25 Gauge, 1&#34;, Stainless steel, PTFE lined.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Plastic dispensing bottle for acrylic cement</td>
			<td style="width:133px;">McMaster Carr</td>
			<td style="width:135px;">7544A67</td>
			<td style="width:345px;">Plastic dispensing bottle, 2-oz size, packs of 5.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Viscous acrylic cement</td>
			<td style="width:133px;">McMaster Carr</td>
			<td style="width:135px;">7515A11</td>
			<td style="width:345px;">Scigrip plastic pipe cement. Medium-bodied acrylic cement to seal in any gaps within PBR body.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">PG-13.5 thread tap</td>
			<td style="width:133px;">McMaster Carr</td>
			<td style="width:135px;">2485A14</td>
			<td style="width:345px;">Can be used to help secure pH electrode to lid (if applicable).</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">PBR and lid</td>
			<td style="width:133px;">NCSU Precision Machine Shop</td>
			<td style="width:135px;">Karam Algae 3.2L Reactor Revision E</td>
			<td style="width:345px;">This machine shop is open to public for business. Contact shop manager.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">pH sensor</td>
			<td style="width:133px;">Hamilton</td>
			<td style="width:135px;">238643</td>
			<td style="width:345px;">EasyFerm Plus 120, autoclavable, millivolt output.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Light sensor</td>
			<td style="width:133px;">Apogee Instruments</td>
			<td style="width:135px;">SQ-225</td>
			<td style="width:345px;">Amplified 0-5 volt electric calibration quantum sensor, water-proof.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">Temperature sensor</td>
			<td style="width:133px;">LabJack</td>
			<td style="width:135px;">EI1034</td>
			<td style="width:345px;">Stainless steel, water-proof temperature sensor.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:169px;">pH transmitter wire with BNC end</td>
			<td style="width:133px;">Sigma-Aldrich</td>
			<td style="width:135px;">HAM355173-1EA</td>
			<td style="width:345px;">This wire will vary with type of pH probe. Make sure wire is compatible with pH probe and has BNC connector end.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Unity gain pre-amplifier</td>
			<td style="width:133px;">Omega Engineering</td>
			<td style="width:135px;">PHTX-21</td>
			<td style="width:345px;">Signal processing amplifier for pH electrode needed for high-impedance pH readings.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Coaxial adapter, BNC female-to-binding post</td>
			<td style="width:133px;">Amazon</td>
			<td style="width:135px;">SMAKN B00NGD5K80</td>
			<td style="width:345px;">For connecting pH signal from pre-amplifier to microcontroller.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">Capacitor (1000 uF)</td>
			<td style="width:133px;">Amazon</td>
			<td style="width:135px;">Nichicon BCBI4950</td>
			<td style="width:345px;">For low-pass filter.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">Resistor (1000 ohm)</td>
			<td style="width:133px;">Radio Shack</td>
			<td style="width:135px;">2711321</td>
			<td style="width:345px;">For low-pass filter.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Hookup wire</td>
			<td style="width:133px;">RadioShack</td>
			<td style="width:135px;">2781222</td>
			<td style="width:345px;">For making low-pass filters, connecting sensors to microcontroller, and wiring motor.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">Heat shrink tubing</td>
			<td style="width:133px;">RadioShack</td>
			<td style="width:135px;">2781611</td>
			<td style="width:345px;">For low-pass filter assembly.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:169px;">Data acquisition and control unit</td>
			<td style="width:133px;">LabJack</td>
			<td style="width:135px;">LabJack U6</td>
			<td style="width:345px;">To process electrical signal from sensors and communicate with data acquisition and control software.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:169px;">DAQFactory data acquisition software</td>
			<td style="width:133px;">DAQFactory</td>
			<td style="width:135px;">DAQFactory Express Release 5.87c Build: 2050</td>
			<td style="width:345px;">Free to download, for up to 10 channels.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">Mini DC-gearmotor</td>
			<td style="width:133px;">McMaster Carr</td>
			<td style="width:135px;">6331K31</td>
			<td style="width:345px;">Motor for mixer impeller.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">Impeller and shaft</td>
			<td style="width:133px;">N/A</td>
			<td style="width:135px;">N/A</td>
			<td style="width:345px;">Email authors for 3D files.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:169px;">Variable DC power supply</td>
			<td style="width:133px;">Amazon</td>
			<td style="width:135px;">Tekpower HY1803D</td>
			<td style="width:345px;">Variable DC power supply, 0-18V @ 0-3A.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:169px;">Grow Lamp</td>
			<td style="width:133px;">HydroGrow</td>
			<td style="width:135px;">SOL-1</td>
			<td style="width:345px;">This exact model is no longer available.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:169px;">Incubator</td>
			<td style="width:133px;">Thermo Scientific</td>
			<td style="width:135px;">Precision Model 818</td>
			<td style="width:345px;">This particular incubator can withstand an internal heat source since this unit&#39;s cooling compressors run non-stop regardless of temperature setting.</td>
		</tr>
	</tbody>
</table>

construction and setup of a bench-scale algal photosynthetic bioreactor with temperature, light, and ph monitoring for kinetic growth tests

用于微藻培养的光合生物反应器（PBR）的最佳设计和操作对于改善微藻类生物燃料生产的环境和经济性能至关重要。在不同条件下估计微藻生长的模型有助于优化PBR设计和操作。为了有效，必须准确确定这些模型中使用的生长参数。藻类生长实验通常受到培养环境的动态性质的限制，需要控制系统来准确地确定动力学参数。实施受控批量实验的第一步是实时数据采集和监测。该协议概述了可用于进行微藻生长实验的台式光合生物反应器的装配和操作的过程。该协议描述了如何从亚克力尺寸和组装平板，台式规模的PBR。它还详细说明了如何配置使用数据采集和控制单元，模拟传感器和开源数据采集软件，连续进行pH，光和温度监测的PBR。

来自该实时监测系统的数据显示了台式PBR中藻类的动态培养环境，并突出了对系统进行监测和控制的需要。记录的温度数据（ 图6 ）显示了如何照明，孵化器空气温度和与藻类生长相关的能量耗散可以改变PBR内的温度，以及如何根据需要使用实时数据来调节孵化器温度控制。 在实验过程中测量的光进一步强调了这种不断增长的环境的动态性质。 如图7所示 ，作为光合光子通量密度（PPFD;μE-m -2 s -1 ）测量的光传感器读数在加入藻类之前为〜100PPFD，立即下降到85PPFD af接着用藻类培养物接种反应器。光照强度在第7天持续下降到5PPFD以下。光照强度的下降是由于生物量和细胞数量的增加，和/或增加叶绿素含量的吸收，表明藻类在第7天活跃，尽管低光照水平。需要额外的生物测量来进一步推断。 连续记录的pH数据显示，总体而言，使用实施的pH控制算法在该实验过程中充分控制pH值（ 图8 ）。这些数据显示了每分钟读数和小时平均值，显示了关于培养藻类和实时监控pH的几个关键点。首先，在用藻类接种PBR之后，pH增加到高于所需设定点7.6。预期这种变化，因为添加到PBR中的培养种子有apH值高于设定值，因为用于培养接种物的烧瓶不进行pH控制。其次，这个实时数据显示了pH电极对外部电气噪声的敏感程度。这种灵敏度由第1天和第2天之间的电极值急剧上升所引起。这些pH值的突然变化很可能是由来自相邻实验装置的电磁阀的电噪声引起的。这种电气干扰过早地触发了pH控制算法，以将CO 2注入到PBR中。因此，pH降低到所需的设定点以下。 pH电极的灵敏度可能导致极端的异常值，并且可能会破坏控制系统。 <img alt="图3" class="xfigimg" src="/files/ftp_upload/55545/55545fig3.jpg" /> 图3:pH响应和校准示例图。 （ a ）实施例的响应图e pH传感器（ b ）pH传感器的示例校准图，用于转换的方程式。回归分析显示95％置信区间。误差条不可见（标准误差小于0.03％）。这些图表显示pH传感器正确连接，其信号非常稳定。 <a href="https://ecsource-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55545/55545fig3large.jpg" target="_blank">请点击此处查看此图的较大版本。</a> <img alt="图6" class="xfigimg" src="/files/ftp_upload/55545/55545fig6.jpg" /> 图6:7天实验期间PBR内的温度测量。深蓝色点表示传感器数据的1小时平均值，浅蓝色点表示在1分钟内获取的传感器读数（采集时间为1秒，平均长度为60），并使用制造商提供的转换因子转换为温度。黑色箭头<html>当调节培养箱温度设置以维持培养温度在25℃左右时（这个所需的设定点用红色，虚线表示）。温度波动是由于藻类生长和培养箱温度的变化。 <a href="https://ecsource-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55545/55545fig6large.jpg" target="_blank">请点击此处查看此图的较大版本。</a> <img alt="图7" class="xfigimg" src="/files/ftp_upload/55545/55545fig7.jpg" /> 图7:7天实验期间PBR内的光测量。深蓝色点表示传感器数据的1小时平均值，浅蓝色点表示在1分钟内获取的传感器读数（采集时间为1秒，平均长度为60），并使用默认光传感器出厂校准值转换为PPFD。"&gt;请点击此处查看此图的较大版本。 <img alt="图8" class="xfigimg" src="/files/ftp_upload/55545/55545fig8.jpg" /> 图8:7天实验期间PBR内的pH测量。深蓝色点表示传感器数据的1小时平均值，浅蓝色点表示每1分钟记录的传感器读数（采集时间为0.1秒，平均长度为600），并使用通过校准建立的转换方程转换为pH。使用99％CO 2气体注入，将pH值保持在7.6至7.5之间。红色的虚线表示所需的pH范围。 <a href="https://ecsource-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/55545/55545fig8large.jpg" target="_blank">请点击此处查看此图的较大版本。</a>

Watch this Scientific Journal Video about Construction and Setup of a Bench-scale Algal Photosynthetic Bioreactor with Temperature, Light, and pH Monitoring for Kinetic Growth Tests at JoVE.com

Construction and Setup of a Bench-scale Algal Photosynthetic Bioreactor with Temperature, Light, and pH Monitoring for Kinetic Growth Tests

本文介绍了可以与其他方法一起使用的台式光合生物反应器的装配过程和操作，以估计相关的动力学生长参数。该系统使用传感器，数据采集和控制单元以及开源数据采集软件连续监控pH，光和温度。

用于动态生长测试的温度，光照和pH监测的台式藻类光合生物反应器的构建和设置

The optimal design and operation of photosynthetic bioreactors (PBRs) for microalgal cultivation is essential for improving the environmental and economic ...

construction-setup-bench-scale-algal-photosynthetic-bioreactor-with

Nanyang Technological University

Research

JoVE Journal

Bioengineering

16.0K 次观看.  North Carolina State University. 本文介绍了可以与其他方法一起使用的台式光合生物反应器的装配过程和操作，以估计相关的动力学生长参数。该系统使用传感器，数据采集和控制单元以及开源数据采集软件连续监控pH，光和温度。 

视频: Construction and Setup of a Bench-scale Algal Photosynthetic Bioreactor with Temperature, Light, and pH Monitoring for Kinetic Growth Tests

Gold Nanostar Synthesis with a Silver Seed Mediated Growth Method

The physical, chemical and optical properties of nano-scale colloids depend on their material composition, size and shape 1-5. There is a great interest in using nano-colloids for photo-thermal ablation, drug delivery and many other biomedical applications 6. Gold is particularly used because of its low toxicity 7-9. A property of metal nano-colloids is that they can have a strong surface plasmon resonance 10. The peak of the surface plasmon resonance mode depends on the structure and composition of the metal nano-colloids. Since the surface plasmon resonance mode is stimulated with light there is a need to have the peak absorbance in the near infrared where biological tissue transmissivity is maximal 11, 12.
We present a method to synthesize star shaped colloidal gold, also known as star shaped nanoparticles 13-15 or nanostars 16. This method is based on a solution containing silver seeds that are used as the nucleating agent for anisotropic growth of gold colloids 17-22. Scanning electron microscopy (SEM) analysis of the resulting gold colloid showed that 70 % of the nanostructures were nanostars. The other 30 % of the particles were amorphous clusters of decahedra and rhomboids. The absorbance peak of the nanostars was detected to be in the near infrared (840 nm). Thus, our method produces gold nanostars suitable for biomedical applications, particularly for photo-thermal ablation.

We synthesized star shaped gold nanostars using a silver seed mediated growth method. The diameter of the nanostars ranges from 200 to 300 nm and the number of tips vary from 7 to 10. The nanoparticles have a broad surface plasmon resonance mode centered in the near infrared.

黄金Nanostar合成银种子介导的生长方法

The physical, chemical and optical properties of nano-scale colloids depend on their material composition, size and shape 1-5. There is a great interest ...

科研

生物工程

Mass Cytometry: Protocol for Daily Tuning and Running Cell Samples on a CyTOF Mass Cytometer

In recent years, the rapid analysis of single cells has commonly been performed using flow cytometry and fluorescently-labeled antibodies. However, the issue of spectral overlap of fluorophore emissions has limited the number of simultaneous probes. In contrast, the new CyTOF mass cytometer by DVS Sciences couples a liquid single-cell introduction system to an ICP-MS.1 Rather than fluorophores, chelating polymers containing highly-enriched metal isotopes are coupled to antibodies or other specific probes.2-5 Because of the metal purity and mass resolution of the mass cytometer, there is no "spectral overlap" from neighboring isotopes, and therefore no need for compensation matrices. Additionally, due to the use of lanthanide metals, there is no biological background and therefore no equivalent of autofluorescence. With a mass window spanning atomic mass 103-203, theoretically up to 100 labels could be distinguished simultaneously. Currently, more than 35 channels are available using the chelating reagents available from DVS Sciences, allowing unprecedented dissection of the immunological profile of samples.6-7
Disadvantages to mass cytometry include the strict requirement for a separate metal isotope per probe (no equivalent of forward or side scatter), and the fact that it is a destructive technique (no possibility of sorting recovery). The current configuration of the mass cytometer also has a cell transmission rate of only ~25%, thus requiring a higher input number of cells.
Optimal daily performance of the mass cytometer requires several steps. The basic goal of the optimization is to maximize the measured signal intensity of the desired metal isotopes (M) while minimizing the formation of oxides (M+16) that will decrease the M signal intensity and interfere with any desired signal at M+16. The first step is to warm up the machine so a hot, stable ICP plasma has been established. Second, the settings for current and make-up gas flow rate must be optimized on a daily basis. During sample collection, the maximum cell event rate is limited by detector efficiency and processing speed to 1000 cells/sec. However, depending on the sample quality, a slower cell event rate (300-500 cells/sec) is usually desirable to allow better resolution between cells events and thus maximize intact singlets over doublets and debris. Finally, adequate cleaning of the machine at the end of the day helps minimize background signal due to free metal.

The steps necessary for daily tuning and optimization of the performance of a CyTOF mass cytometer are described. Comments on optimal sample preparation and flow rate are discussed

质量流式细胞术:协议，每日调整和执行细胞的样品上一个CyTOF的质量流式细胞仪

In recent years, the rapid analysis of single cells has commonly been performed using flow cytometry and fluorescently-labeled antibodies. However, the ...

Design of a Biaxial Mechanical Loading Bioreactor for Tissue Engineering

We designed a loading device that is capable of applying uniaxial or biaxial mechanical strain to a tissue engineered biocomposites fabricated for transplantation. While the device primarily functions as a bioreactor that mimics the native mechanical strains, it is also outfitted with a load cell for providing force feedback or mechanical testing of the constructs. The device subjects engineered cartilage constructs to biaxial mechanical loading with great precision of loading dose (amplitude and frequency) and is compact enough to fit inside a standard tissue culture incubator. It loads samples directly in a tissue culture plate, and multiple plate sizes are compatible with the system. The device has been designed using components manufactured for precision-guided laser applications. Bi-axial loading is accomplished by two orthogonal stages. The stages have a 50 mm travel range and are driven independently by stepper motor actuators, controlled by a closed-loop stepper motor driver that features micro-stepping capabilities, enabling step sizes of less than 50 nm. A polysulfone loading platen is coupled to the bi-axial moving platform. Movements of the stages are controlled by Thor-labs Advanced Positioning Technology (APT) software. The stepper motor driver is used with the software to adjust load parameters of frequency and amplitude of both shear and compression independently and simultaneously. Positional feedback is provided by linear optical encoders that have a bidirectional repeatability of 0.1 &mu;m and a resolution of 20 nm, translating to a positional accuracy of less than 3 &mu;m over the full 50 mm of travel. These encoders provide the necessary position feedback to the drive electronics to ensure true nanopositioning capabilities. In order to provide the force feedback to detect contact and evaluate loading responses, a precision miniature load cell is positioned between the loading platen and the moving platform. The load cell has high accuracies of 0.15% to 0.25% full scale.

We designed a novel mechanical loading bioreactor that can apply uniaxial or biaxial mechanical strain to a cartilage biocomposite prior to transplantation into an articular cartilage defect.

组织工程生物反应器的双轴机械装载设计

We  designed a loading device that is capable of applying uniaxial or biaxial  mechanical strain to a tissue engineered biocomposites fabricated for  ...

Microfluidic Picoliter Bioreactor for Microbial Single-cell Analysis: Fabrication, System Setup, and Operation

In this protocol the fabrication, experimental setup and basic operation of the recently introduced microfluidic picoliter bioreactor (PLBR) is described in detail. The PLBR can be utilized for the analysis of single bacteria and microcolonies to investigate biotechnological and microbiological related questions concerning, e.g.&#160;cell growth, morphology, stress response, and metabolite or protein production on single-cell level. The device features continuous media flow enabling constant environmental conditions for perturbation studies, but in addition allows fast medium changes as well as oscillating conditions to mimic any desired environmental situation. To fabricate the single use devices, a silicon wafer containing sub micrometer sized SU-8 structures served as the replication mold for rapid polydimethylsiloxane casting. Chips were cut, assembled, connected, and set up onto a high resolution and fully automated microscope suited for time-lapse imaging, a powerful tool for spatio-temporal cell analysis. Here, the biotechnological platform organism Corynebacterium glutamicum&#160;was seeded into the PLBR and cell growth and intracellular fluorescence were followed over several hours unraveling time dependent population heterogeneity on single-cell level, not possible with conventional analysis methods such as flow cytometry. Besides insights into device fabrication, furthermore, the preparation of the preculture, loading, trapping of bacteria, and the PLBR cultivation of single cells and colonies is demonstrated. These devices will add a new dimension in microbiological research to analyze time dependent phenomena of single bacteria under tight environmental control. Due to the simple and relatively short fabrication process the technology can be easily adapted at any microfluidics lab and simply tailored towards specific needs.

In this protocol the fabrication, setup and basic operation of a microfluidic picoliter bioreactor (PLBR) for single-cell analysis of prokaryotic microorganisms is introduced. Industrially relevant microorganisms were analyzed as proof of principle allowing insights into growth rate, morphology, and phenotypic heterogeneity over certain time periods, hardly possible with conventional methods.

In this protocol the fabrication, experimental setup and basic operation of the recently introduced microfluidic picoliter bioreactor (PLBR) is described ...

Operation of a Benchtop Bioreactor

Fermentation systems are used to provide an optimal growth environment for many different types of cell cultures. The ability afforded by fermentors to carefully control temperature, pH, and dissolved oxygen concentrations in particular makes them essential to efficient large scale growth and expression of fermentation products. This video will briefly describe the advantages of the fermentor over the shake flask. It will also identify key components of a typical benchtop fermentation system and give basic instruction on setup of the vessel and calibration of its probes. The viewer will be familiarized with the sterilization process and shown how to inoculate the growth medium in the vessel with culture. Basic concepts of operation, sampling, and harvesting will also be demonstrated. Simple data analysis and system cleanup will also be discussed.

Fermentors are used to increase culture yield and productivity of bioengineered cells. After screening multiple microbial or animal cell culture candidates in shake flasks, the next logical step is to increase the selected culture&#x2019;s biomass with the fermentor. This video demonstrates the setup and operation of a typical benchtop bioreactor system.

一个台式生物反应器的操作

Fermentation systems are used to provide an optimal growth environment for many different types of cell cultures. The ability afforded by fermentors to ...

Construction and Characterization of a Novel Vocal Fold Bioreactor

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To emulate the dynamic environment of vocal folds, a novel vocal fold bioreactor capable of producing vibratory stimulations at fundamental phonation frequencies is constructed and characterized. The device is composed of a function generator, a power amplifier, a speaker selector and parallel vibration chambers. Individual vibration chambers are created by sandwiching a custom-made silicone membrane between a pair of acrylic blocks. The silicone membrane not only serves as the bottom of the chamber but also provides a mechanism for securing the cell-laden scaffold. Vibration signals, generated by a speaker mounted underneath the bottom acrylic block, are transmitted to the membrane aerodynamically by the oscillating air. Eight identical vibration modules, fixed on two stationary metal bars, are housed in an anti-humidity chamber for long-term operation in a cell culture incubator. The vibration characteristics of the vocal fold bioreactor are analyzed non-destructively using a Laser Doppler Vibrometer (LDV). The utility of the dynamic culture device is demonstrated by culturing cellular constructs in the presence of 200-Hz sinusoidal vibrations with a mid-membrane displacement of 40 &#181;m. Mesenchymal stem cells cultured in the bioreactor respond to the vibratory signals by altering the synthesis and degradation of vocal fold-relevant, extracellular matrix components. The novel bioreactor system presented herein offers an excellent in vitro platform for studying vibration-induced mechanotransduction and for the engineering of functional vocal fold tissues.

A novel vocal fold bioreactor capable of delivering physiologically relevant, vibratory stimulation to cultured cells is constructed and characterized. This dynamic culture device, when combined with a fibrous poly(&#949;-caprolactone) scaffold, creates a vocal fold-mimetic environment that modulates the behaviors of mesenchymal stem cells.

一种新型声带生物反应器的构建与鉴定

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To ...

Light-mediated Formation and Patterning of Hydrogels for Cell Culture Applications

Click chemistries have been investigated for use in numerous biomaterials applications, including drug delivery, tissue engineering, and cell culture. In particular, light-mediated click reactions, such as photoinitiated thiol&#8722;ene and thiol&#8722;yne reactions, afford spatiotemporal control over material properties and allow the design of systems with a high degree of user-directed property control. Fabrication and modification of hydrogel-based biomaterials using the precision afforded by light and the versatility offered by these thiol&#8722;X photoclick chemistries are of growing interest, particularly for the culture of cells within well-defined, biomimetic microenvironments. Here, we describe methods for the photoencapsulation of cells and subsequent photopatterning of biochemical cues within hydrogel matrices using versatile and modular building blocks polymerized by a thiol&#8722;ene photoclick reaction. Specifically, an approach is presented for constructing hydrogels from allyloxycarbonyl (Alloc)-functionalized peptide crosslinks and pendant peptide moieties and thiol-functionalized poly(ethylene glycol) (PEG) that rapidly polymerize in the presence of lithium acylphosphinate photoinitiator and cytocompatible doses of long wavelength ultraviolet (UV) light. Facile techniques to visualize photopatterning and quantify the concentration of peptides added are described. Additionally, methods are established for encapsulating cells, specifically human mesenchymal stem cells, and determining their viability and activity. While the formation and initial patterning of thiol-alloc hydrogels are shown here, these techniques broadly may be applied to a number of other light and radical-initiated material systems (e.g., thiol-norbornene, thiol-acrylate) to generate patterned substrates.

We describe a sequential process for light-mediated formation and subsequent biochemical patterning of synthetic hydrogel matrices for three-dimensional cell culture applications. The construction and modification of hydrogels with cytocompatible photoclick chemistry is demonstrated. Additionally, facile techniques to quantify and observe patterns and determine cell viability within these hydrogels are presented.

用于细胞培养应用光介导的形成和构图的水凝胶

Click chemistries have been investigated for use in numerous biomaterials applications, including drug delivery, tissue engineering, and cell culture. In ...

Optimized Setup and Protocol for Magnetic Domain Imaging with In Situ Hysteresis Measurement

This paper elaborates the sample preparation protocols required to obtain optimal domain patterns using the Bitter method, focusing on the extra steps compared to standard metallographic sample preparation procedures. The paper proposes a novel bespoke rig for dynamic domain imaging with in situ BH (magnetic hysteresis) measurements and elaborates the protocols for the sensor preparation and the use of the rig to ensure accurate BH measurement. The protocols for static and ordinary dynamic domain imaging (without in situ BH measurements) are also presented. The reported method takes advantage of the convenience and high sensitivity of the traditional Bitter method and enables in situ BH measurement without interrupting or interfering with the domain wall movement processes. This facilitates establishing a direct and quantitative link between the domain wall movement processes&#8211;microstructural feature interactions in ferritic steels with their BH loops. This method is anticipated to become a useful tool for the fundamental study of microstructure&#8211;magnetic property relationships in steels and to help interpret the electromagnetic sensor signals for non-destructive evaluation of steel microstructures.

Optimized Setup and Protocol for Magnetic Domain Imaging with In Situ Hysteresis Measurement

This paper elaborates the sample and sensor preparation procedures and the protocols for using the test rig particularly for dynamic domain imaging with in situ BH measurements in order to achieve optimal domain pattern quality and accurate BH measurements.

用原位滞后测量进行磁畴成像的优化设置和协议

This paper elaborates the sample preparation protocols required to obtain optimal domain patterns using the Bitter method, focusing on the extra steps ...

Construction of a Multilayered Mesenchymal Stem Cell Sheet with a 3D Dynamic Culture System

Stem cell therapy shows a promising future in regenerating injured organ and tissues, and the cell sheet technique has been developed to improve the low cell retention and poor survival within the target zone. However, during the in vitro construction process, a solution for maintaining stem cell bioactivity and increasing the cell amount within the cell sheet is urgently needed. Here, this protocol presents a method for constructing a multilayered cell sheet with favorable stem cell bioactivity and optimal operability. Decellularized porcine pericardium (DPP) is prepared by phospholipase A2 (PLA2) decellularization method as the cell sheet scaffold, and rat bone marrow mesenchymal stem cells (BMSCs) are isolated and expanded as the seeded cells. The temporary multilayered cell sheet structure is constructed by using RAD16-I peptide hydrogel. Finally, the cell sheet is cultured with a dynamic perfusion system to stabilize the three-dimensional (3D) structure, and the cell sheet could be obtained following a 48-hour culture in vitro. This protocol provides an efficient and feasible method for constructing a multilayered stem cell sheet, and the cell sheet could be developed as a favorable stem cell therapy product in the future.

This article provides an efficient and feasible method for constructing multilayered stem cell sheets with favorable stem cell property.

3D 动态培养系统构建多层间充质干细胞片材

Stem cell therapy shows a promising future in regenerating injured organ and tissues, and the cell sheet technique has been developed to improve the low ...

Construction of a Wireless-Enabled Endoscopically Implantable Sensor for pH Monitoring with Zero-Bias Schottky Diode-based Receiver

Ambulatory pH monitoring of pathological reflux is an opportunity to observe the relationship between symptoms and exposure of the esophagus to acidic or non-acidic refluxate. This paper describes a method for the development, manufacturing, and implantation of a miniature wireless-enabled pH sensor. The sensor is designed to be implanted endoscopically with a single hemostatic clip. A fully passive rectenna-based receiver based on a zero-bias Schottky diode is also constructed and tested. To construct the device, a two-layer printed circuit board and off-the-shelf components were used. A miniature microcontroller with integrated analog peripherals is used as an analog front end for the ion-sensitive field-effect transistor (ISFET) sensor and to generate a digital signal which is transmitted with an amplitude shift keying transmitter chip. The device is powered by two primary alkaline cells. The implantable device has a total volume of 0.6 cm3 and a weight of 1.2 grams, and its performance was verified in an ex vivo model (porcine esophagus and stomach). Next, a small footprint passive rectenna-based receiver which can be easily integrated either into an external receiver or the implantable neurostimulator, was constructed and proven to receive the RF signal from the implant when in proximity (20 cm) to it. The small size of the sensor provides continuous pH monitoring with minimal obstruction of the esophagus. The sensor could be used in routine clinical practice for 24/96 h esophageal pH monitoring without the need to insert a nasal catheter. The &#34;zero-power&#34; nature of the receiver also enables the use of the sensor for automatic in-vivo calibration of miniature lower esophageal sphincter neurostimulation devices. An active sensor-based control enables the development of advanced algorithms to minimize the used energy to achieve a desirable clinical outcome. One of the examples of such an algorithm would be a closed-loop system for on-demand neurostimulation therapy of gastroesophageal reflux disease (GERD).

The manuscript presents a miniature implantable pH sensor with ASK modulated wireless output together with a fully passive receiver circuit based on zero-bias Schottky diodes. This solution can be used as a basis in the development of in vivo calibrated electrostimulation therapy devices and for ambulatory pH monitoring.

构建一种无线内窥镜植入式传感器，用于使用基于零偏置肖特基二极管的接收器进行pH监测

Ambulatory pH monitoring of pathological reflux is an opportunity to observe the relationship between symptoms and exposure of the esophagus to acidic or ...