涉及无脊椎动物的协议免于布坦坦研究所动物使用和护理委员会的批准。然而，这里描述的蝎子的维护遵循道德参数，并且根据物种的需要尊重动物福利。
1. 住房
<ol><li>将蝎子饲养在公共聚丙烯容器中，作为围栏（高 35 厘米，宽 35.5 厘米，长 72 厘米），最多可容纳 350 只动物。</li>
	<li>将饲养动物围栏的房间保持在24°C（±0.5°C）的受控温度下，在灯打开10小时和灯关闭14小时的光循环中。</li>
	<li>在每个容器的外壳地板（底部）上铺上牛皮纸，在四面八方的边缘用遮蔽胶带固定，这样动物就不会进入纸下面（图3A）。</li>
	<li>使用四个纸板托盘（蛋托盘）作为垂直基材，堆叠在容器的一侧，每个托盘之间放置一块硬纸板（图3B，C）。</li>
	<li>在外壳的另一侧放置一个浅聚丙烯托盘（水托盘），其中包含浸泡的棉花作为动物的水源（图3B）。</li>
	<li>保持容器不带盖以增加气流。在外壳内表面的顶部和上角放置一条约10厘米的自粘塑料胶带，以使墙壁足够光滑，以防止动物从容器中逸出（图3A）。</li>
</ol><img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/65737/65737fig03.jpg"> 图 3：外壳和外壳准备 。 （A） 箭头表示底部的牛皮纸，边缘用遮蔽胶带固定。虚线表示放置在外壳内表面顶部的自粘塑料胶带。（B） 垂直基材（堆在容器一侧的蛋托）和放置在另一侧的浅聚丙烯托盘（水托盘）。（C） 垂直基材细节：在两个托盘之间用硬纸板起球的蛋托。（D）层层放置的纸板和蛋托，模拟 了沙氏毛蛾 的自然黑暗和潮湿栖息地。 （E） 饲养在基质中的动物。 <a href="https://www.jove.com/files/ftp_upload/65737/65737fig03large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
2. 卫生习惯
注意： 卫生程序分为两个维护程序：完全维护和部分维护。第一个是在喂养动物后2天或选择围栏进行毒液提取时进行的。当动物没有被喂食时，进行部分维护。
<ol><li>全面维护<ol><li>如第 1 节所述，准备一个带有新基材（蛋托、纸板和牛皮纸）和干净水托的干净外壳。</li>
		<li>使用长镊子，从外壳上取下水盘。如果水盘上有蝎子，请将它们取出并放入干净的外壳中。</li>
		<li>对所有蛋托和纸板重复步骤 2.1.2 中描述的相同步骤，取出所有蝎子并将它们放入干净的外壳中。</li>
		<li>取下托盘后，取出所有留在外壳地板上的蝎子，并将它们放入干净的外壳中。请务必考虑任何尸体并妥善丢弃它们。收集所有剩余的活猎物（昆虫）并将它们从围栏中取出。</li>
		<li>从外壳中取出所有一次性基材（蛋托、纸板和牛皮纸）并正确丢弃。将可重复使用的物品（水盘和聚丙烯容器）送去进行卫生。 注意： 可重复使用物品的卫生是用 0.5% 次氯酸盐和中性清洁剂进行的。使用任何卫生产品或溶液后，必须用流水彻底冲洗材料。</li>
	</ol></li>
	<li>部分维护 注意： 部分维护包括检查所有基材（蛋托和牛皮纸），目的是去除死蝎子尸体和其他不需要的碎屑，但不要丢弃基材或更换外壳。<ol><li>使用长镊子轻轻提起每个蛋托并清除废物进行检查。</li>
		<li>如果需要，用干净的蛋托更换一个或多个蛋托。</li>
		<li>使用长镊子用干净的水盘完全更换水盘。将水盘上的任何蝎子转移到外壳另一部分的蛋托上。</li>
	</ol></li>
</ol>3. 喂食
<ol><li>根据毒液提取时间表，每15天喂一次动物。喂养在毒液提取前 7 天和提取毒液后 7 天挤奶的动物。</li>
	<li>对于喂食，交替供应蟑螂（Phoetalia pallida）和蟋蟀（Gryllus sp.）。</li>
	<li>根据动物的能量摄入量计算食物量;为围栏内的每 1 或 3 只蝎子提供 5 个猎物。</li>
	<li>计算出的猎物数量并将它们活着放入围栏内，在完全维护之前将它们放置 2 天（第 2.1 节）。</li>
	<li>提供蟑螂时，在外壳顶部周围涂抹碳酸钙和酒精溶液。该解决方案导致蟑螂打滑，无法爬上围栏的侧面，从而防止昆虫逃逸。</li>
</ol>4.毒液提取
注意：毒液提取程序中使用的动物是成虫或成虫前。由于难以确定巴西黄蝎子的完全性成熟（成年期），因此确定体长为 5 至 7 厘米的动物有资格进行该手术。
<ol><li>每 2-3 个月给每头动物挤奶一次。</li>
	<li>在提取毒液的当天，计算围栏内所有活的动物，并使用长镊子将它们转移到玻璃容器中（图4）。</li>
	<li>准备和布置气流柜工作台面顶部要使用的所有材料和设备。将电刺激器电源线插入插座，然后打开电刺激器和气流柜（图 2B）。<ol><li>使用以下电刺激器参数设置：电位（电极之间的电位差）：540 mV，应力（电脉冲持续时间）：2 ms，作用（间歇性以调节使用的效力，这将减少 telson 有机组织的加热）：70%，循环（调整循环重复）：1 s，强度（通过动物肌肉循环的电流）： 0.75毫安。</li>
	</ol></li>
	<li>按照以下步骤手动约束每只动物，使 telson 静止不动：<ol><li>用直镊子握住动物的变形体（图5A）。</li>
		<li>用弯曲的镊子固定telson（图5B）。</li>
		<li>用手束缚telson（图5C，D）。</li>
	</ol></li>
	<li>约束后，将telson与电刺激器导体直接接触，无需使用导电溶液。电击导致毒液腺的肌肉不自主收缩，随后毒液喷射。</li>
	<li>将毒液滴直接收集到靠近电刺激器导体的塑料微管中（图6）。</li>
	<li>接下来，按照以下步骤将动物放回玻璃容器中：<ol><li>将动物的端子从手转移到弯曲的镊子上。</li>
		<li>将动物轻轻地放在玻璃容器的底部。</li>
	</ol></li>
	<li>对玻璃杯中的每只动物执行步骤4.4-4.7。</li>
	<li>在玻璃杯中挤奶的所有动物都挤完奶后，将它们放回围栏内。</li>
	<li>将收集的毒液储存在-20°C，直到冻干过程。</li>
</ol><img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/65737/65737fig04.jpg"> 图4：玻璃容器中的蝎子。 <a href="https://www.jove.com/files/ftp_upload/65737/65737fig04large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/65737/65737fig05.jpg"> 图 5：毒液提取的限制程序 。 （A） 用直镊子（黑色箭头）束缚的 Metassoma。（B） Telson用弯曲的镊子固定（黑色箭头）。（C） 将受约束的端子从镊子转移到手上。（D） 特尔森被手束缚。建议徒手处理动物，因为当使用手套时，蝎子会用爪子抓住材料，因此在手术结束时难以释放它们，并大大增加了发生事故的风险。 <a href="https://www.jove.com/files/ftp_upload/65737/65737fig05large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/65737/65737fig06.jpg"> 图 6：毒液提取。 放置在靠近电刺激器（黄色箭头）的塑料微管（白色箭头）。在手术过程中，毒液滴被直接收集到保持在室温的管中。 <a href="https://www.jove.com/files/ftp_upload/65737/65737fig06large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>

饲养巴西黄蝎子提取毒液

<ol>
	<li>Polis, G. 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> The biology of scorpions. , (1990).</li><li>Lacerda, A. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scorpion+envenomation+in+the+state+of+S&#227;o+Paulo,+Brazil:+Spatiotemporal+analysis+of+a+growing+public+health+concern.">Scorpion envenomation in the state of S&#227;o Paulo, Brazil: Spatiotemporal analysis of a growing public health concern.</a> PLoS One. 17 (4), 0266138 (2022).</li><li>Reckziegel, G. C., Pinto, V. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scorpionism+in+Brazil+in+the+years+2000+to+2012.">Scorpionism in Brazil in the years 2000 to 2012.</a> Journal of Venomous Animals and Toxins Including Tropical Diseases. 20, 46 (2014).</li><li>Guerra-Duarte, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scorpion+envenomation+in+Brazil:+Current+scenario+and+perspectives+for+containing+an+increasing+health+problem.">Scorpion envenomation in Brazil: Current scenario and perspectives for containing an increasing health problem.</a> PLoS Neglected Tropical Diseases. 17 (2), 0011069 (2023).</li><li>Pimenta, R. J. G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selected+to+survive+and+kill:+Tityus+serrulatus,+the+Brazilian+yellow+scorpion.">Selected to survive and kill: Tityus serrulatus, the Brazilian yellow scorpion.</a> PLoS One. 14 (4), e0214075 (2019).</li><li>Lacerda, A. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detection+of+areas+vulnerable+to+scorpionism+and+its+association+with+environmental+factors+in+S&#227;o+Paulo,+Brazil.">Detection of areas vulnerable to scorpionism and its association with environmental factors in S&#227;o Paulo, Brazil.</a> Acta Tropica. 230, 106390 (2022).</li><li>Torrez, P. P. Q., Dourado, F. S., Bertani, R., Cupo, P., Fran&#231;a, F. O. d. e. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scorpionism+in+Brazil:+exponential+growth+of+accidents+and+deaths+from+scorpion+stings.">Scorpionism in Brazil: exponential growth of accidents and deaths from scorpion stings.</a> Journal of the Brazilian Society of Tropical Medicine. 52, 20180350 (2019).</li><li>Amado, T. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vulnerable+areas+to+accidents+with+scorpions+in+Brazil.">Vulnerable areas to accidents with scorpions in Brazil.</a> Tropical Medicine &amp; International Health. 26 (5), 13561 (2021).</li><li>Brasil, Minist&#233;rioda da Sa&#250;de, Funda&#231;&#227;o Nacional de Sa&#250;de. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Manual de diagn&#243;stico e tratamento de acidentes por animais pe&#231;onhentos. , (2001).</li><li>Candido, D. M., Lucas, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maintenance+of+scorpions+of+the+genus+Tityus+Koch+(Scorpiones,+Buthidae)+for+venom+obtention+at+Instituto+Butantan,+S&#227;o+Paulo,+Brazil.">Maintenance of scorpions of the genus Tityus Koch (Scorpiones, Buthidae) for venom obtention at Instituto Butantan, S&#227;o Paulo, Brazil.</a> Journal of Venomous Animals and Toxins including Tropical Diseases. 10 (1), 000100007 (2004).</li><li>Lucas, S. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laborat&#243;rio+de+Artr&#243;podes+do+Instituto+Butantan+e+os+aracn&#237;deos+pe&#231;onhentos.">Laborat&#243;rio de Artr&#243;podes do Instituto Butantan e os aracn&#237;deos pe&#231;onhentos.</a> Hist&#243;ria Ci&#234;ncias Sa&#250;de Manguinhos. 10 (3), 000300011 (2003).</li><li>Braga-Pereira, G. F., Santos, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Asexual+reproduction+in+a+sexual+population+of+the+Brazilian+yellow+scorpion+(Tityus+serrulatus,+Buthidae)+as+evidence+of+facultative+parthenogenesis.">Asexual reproduction in a sexual population of the Brazilian yellow scorpion (Tityus serrulatus, Buthidae) as evidence of facultative parthenogenesis.</a> The Journal of Arachnology. 49 (2), (2021).</li></ol>

作者没有要披露的利益冲突。

所描述方法的应用使我们能够保留大量的 锯齿虫 个体，并使我们对每年毒液生产所需的蝎子数量具有一致的可预测性。这样，我们就能够提前提供足够批次的毒液来供应抗蛇毒血清的制造过程。同时，制定预先制定的维护、喂养和毒液提取时间表是活动的重要组成部分，并有助于遵守所述方案。因此，必须建立例程以保持连续生产。
据作者所知，没有科学描述的关于...

蝎子是属于蛛形纲的节肢动物，蝎子目，包括大约 2,621 种 1,2。这些动物的地理范围很广，分布在除南极洲以外的所有大陆1.蝎子引起的毒液毒液每年导致全球数千人发病或死亡3.据估计，2019 年，这些动物每年造成超过 120 万起事故和 3,500 人死亡。在巴西，病例数呈指数级增长，自 2017 年以来每年达到 100,000 多例4、5、6。过去几十年来，巴西观察到不受控制的城市化，没有充分的污水处理和定期收集和处置垃圾，与环境退化和气候变化有关，为入侵蝎子的繁殖提供了条件，如锯齿蝎，增加了与人类的接触，从而导致了有害事故4,7,8.巴西大约有 178 种蝎子，但具有医学重要性的蜇伤是由 Tityus 属引起的，其中四种（T. serrulatus、T. bahiensis、T. stigmurus 和 T. obscurus）是医学问题，其中 Tityus serrulatus 是造成最严重病例和死亡的原因 7,9.
布坦坦研究所是巴西主要的抗蛇毒血清生产实验室，为巴西卫生系统提供约70,000瓶蝎子抗蛇毒血清。简而言之，抗蛇毒血清生产所涉及的步骤包括在马中接种毒液衍生抗原，收集和纯化富抗体血浆，从而产生 5 mL 小瓶蝎子抗蛇毒血清。每瓶抗蛇毒血清至少能中和 1 毫克蝎子毒液。节肢动物生物群是工业中心不可分割的一部分，负责为蝎子抗蛇毒血清提供源材料。
布坦坦研究所的节肢动物生物馆最初被命名为节肢动物实验室，成立于 1967 年。然而，直到 1995 年，实验室才搬到了专门用于饲养有毒节肢动物10,11 的专用设施。当时，蝎子被饲养在一个 12 m2 的房间里，没有控制温度，分布在大约 13 个围栏中，每个围栏最多可容纳 300 只动物10.由于多年来对毒液的需求不断增加，随之而来的需要增加动物数量，以及毒液提取过程的改进，2016 年发生了重大变化。该实验室并入布坦坦研究所的工业中心，并更名为节肢动物生物馆。此外，在 2016 年，Bioterium 搬到了一个新的专用设施。新的蝎子室占地 24 m2，包含大约 48 个聚丙烯围栏，每个围栏最多可容纳 300 只动物，如图 1 所示。这导致总共有 10,000 到 20,000 人被维持在受控圈养条件下，全年都在变化。蝎子在严格的饲养规程下得到精心管理，以确保它们的福利，同时满足高生产要求并遵守动物护理的良好规范和道德标准。为了提取毒液，设计和建造了一个特定的房间，容纳了两个气流柜，如图2A所示。这些机柜作为安全措施，防止技术人员在提取过程中吸入毒液颗粒。技术人员必须配备个人防护装备 （PPE），包括合成围裙和 FFP2 口罩，以避免任何污染并确保他们的安全。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/65737/65737fig01v2.jpg"> 图1：节肢动物生物群的蝎子房间的一般视图。 <a href="https://www.jove.com/files/ftp_upload/65737/65737fig01large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/65737/65737fig02.jpg"> 图 2：工作区 。 （A） 节肢动物生物群的毒液提取室的一般视图。（B） 气流柜工作台面。所有材料和设备都准备好并布置在气流柜工作台面的顶部。黑色箭头表示电刺激器装置，红色箭头表示镊子。 <a href="https://www.jove.com/files/ftp_upload/65737/65737fig02large.jpg" target="_blank">请点击这里查看此图的较大版本.</a>
节肢动物生物群中约90%的 锯齿蝎 子来自不同的地点，由市政卫生人员收集，作为这些动物监测和控制计划的一部分。这些蝎子在城市地区被捕获，然后作为合作计划的一部分被送往布坦坦研究所。 锯齿蝎 是孤雌生殖的，这意味着出生是从未受精卵发育后代的结果，没有异性个体的存在12.由于这种类型的繁殖，一些动物在我们的设施中被圈养出生，并按照如下所述的类似饲养方案进行饲养，直到它们达到成年期。一旦它们达到成虫阶段，它们就会被添加到毒液提取程序中。抵达后，蝎子会接受初步筛选，健康状况良好的蝎子将被饲养在最多可容纳 350 只的集体围栏中，按其来源划分。对于每个围栏，每天填写一份控制表，提供有关喂食日期和提供的猎物类型（蟋蟀或蟑螂）、供水、清洁程序、死亡动物数量以及围栏内剩余活动物数量的数据。
在预先安排的议程中，动物受到电刺激，然后将数百个体的毒液冻干，从而产生成批的冻干毒液。从该工艺中获得的标准化毒液作为制造过程的一部分用于马免疫，并为活性物质和最终产品的质量控制提供参考物质。
世界上很少有设施能够饲养如此多的蝎子，并按照良好的生产规范和动物的道德使用进行大量毒液提取，就像布坦坦研究所的节肢动物生物群所做的那样。因此，我们的目标是描述 锯齿 蝎饲养中使用的蝎子维持方案和毒液提取程序，它们成功地为蝎子抗蛇毒血清的生产提供了必要量的毒液。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Carboard sheet</td>
			<td>NA</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Egg tray</td>
			<td>NA</td>
			<td>NA</td>
			<td>cardboard 36 places egg trays</td>
		</tr>
		<tr>
			<td>Electro stimulator device</td>
			<td>Instituto Butantan</td>
			<td>ART-FV-01</td>
			<td>Internally designed for the venom extraction&#160;</td>
		</tr>
		<tr>
			<td>Kraft paper</td>
			<td>NA</td>
			<td>NA</td>
			<td>regular kraft paper</td>
		</tr>
		<tr>
			<td>Making tape</td>
			<td>NA</td>
			<td>NA</td>
			<td>White masking tape (24 mm x 50 m)</td>
		</tr>
		<tr>
			<td>Polyproplene container</td>
			<td>NA</td>
			<td>NA</td>
			<td>70 L Polypropylene box (H &#8211; 35 cm, W &#8211; 35.5 cm, L - 72 cm)</td>
		</tr>
		<tr>
			<td>Polyproplene tray</td>
			<td>NA</td>
			<td>NA</td>
			<td>Polypropylene tray (H &#8211; 3.0 cm, W &#8211; 18.5 cm, L - 30 cm)</td>
		</tr>
		<tr>
			<td>Self-adhesive plastic</td>
			<td>NA</td>
			<td>NA</td>
			<td></td>
		</tr>
	</tbody>
</table>

captive maintenance and venom extraction of tityus serrulatus (brazilian yellow scorpion) for antivenom production

蝎子毒液是几个热带和亚热带国家的公共卫生问题。 Tityus serrulatus Lutz 和 Mello，1922 年（巴西黄蝎子）每年在巴西造成约 150,000 例毒液中毒病例，其中 10% 需要抗蛇毒血清治疗以逆转危及生命的毒液效应。因此，数以千计的 锯齿锥虫 个体被维持在受控的圈养条件下进行毒液提取，随后用于生产国家供应的蝎子抗蛇毒血清。布坦坦研究所是巴西主要的抗蛇毒血清生产实验室，为巴西卫生系统提供约70,000瓶蝎子抗蛇毒血清。因此，饲养方案和毒液提取方法是成功实现大规模、标准化毒液生产的关键点。本文的目的是描述 锯齿毛虫 饲养的圈养协议，包括饲养程序和毒液提取程序，遵循良好的生产规范，并确保动物福利。这些做法允许饲养多达 20,000 只圈养动物，根据抗蛇毒血清生产需求每月挤奶 3,000 至 5,000 只蝎子，平均达到 90% 的阳性提取率。

Scorpions are arthropods belonging to the class Arachnida, order Scorpiones, which comprises approximately 2,621 species1,2. These animals have a wide geographical range and are present on all continents except Antarctica1. Envenomation caused by scorpions results in the morbidity or death of thousands of people every year globally3. In 2019, it was estimated that there were more than 1.2 million accidents and 3,500 annual deaths caused by these animals. In Brazil, the number of cases has been increasing exponentially, reaching more than 100,000 cases per year since 20174,5,6. The uncontrolled urbanization observed in Brazil in the past decades, without adequate sewage treatment and regular collection and disposal of garbage, associated with environmental degradation and climate changes, has provided conditions for the proliferation of invasive scorpions, such as T. serrulatus, increasing the contact with humans and thus resulting in harmful accidents4,7,8. There are about 178 scorpion species in Brazil, but stings of medical importance are caused by the genus Tityus, with four species (T. serrulatus, T. bahiensis, T. stigmurus, and T. obscurus) being of medical concern, with Tityus serrulatus being responsible for the most severe cases and deaths7,9.
Instituto Butantan is the main antivenom-manufacturing laboratory in Brazil, providing approximately 70,000 vials of scorpion antivenom for the Brazilian health system. Briefly, the steps involved in antivenom manufacturing include the inoculation of venom-derived antigens in equines, the collection and purification of rich-antibodies plasma, resulting in 5 mL vials of scorpion antivenom. Each vial is capable of neutralizing, at least, 1 mg of scorpion venom per mL of antivenom. The Arthropods Bioterium is an integral part of the industrial center, responsible for supplying the source material for the scorpion antivenom.
The Arthropods Bioterium at Instituto Butantan was originally named the Laboratory of Arthropods and was established in 1967. However, it was only in 1995 that the Laboratory moved to an exclusive facility dedicated to housing venomous arthropods10,11. At that time, the scorpions were kept in a room of 12 m2 with no controlled temperature and distributed among approximately 13 enclosures, each housing a maximum of 300 animals10. Due to the increasing demand for venom over the years and the consequent need to increase the number of animals, along with improvements in the venom extraction process, a major change occurred in 2016. The Laboratory was integrated into the industrial center of Instituto Butantan and was renamed the Arthropods Bioterium. Additionally, in 2016, the Bioterium moved to a new purpose-built facility. The new scorpion room covers an area of 24 m2 and contains approximately 48 polypropylene enclosures, each housing up to 300 animals, as shown in Figure 1. This results in a total of 10,000 to 20,000 individuals being maintained under controlled captive conditions, with variations throughout the year. The scorpions are carefully managed under strict husbandry protocols to ensure their welfare while meeting the high production demands and adhering to good practices and ethical standards for animal care. For venom extraction, a specific room was designed and constructed, housing two airflow cabinets as shown in Figure 2A. These cabinets serve as safety measures to prevent technicians from inhaling venom particles during the extraction procedure. Personal protective equipment (PPE) is mandatory for the technicians, including synthetic aprons and FFP2 masks, to avoid any contamination and ensure their safety.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/65737/65737fig01v2.jpg" /> 
Figure 1: General view of the scorpion&#39;s room of the Arthropods Bioterium. <a href="https://www.jove.com/files/ftp_upload/65737/65737fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/65737/65737fig02.jpg" /> 
Figure 2: Working area. (A) General view of the venom extraction room of the Arthropods Bioterium. (B) Airflow cabinet work surface. All the materials and equipment are prepared and arranged on the top of the airflow cabinet work surface. The black arrow indicates the electrostimulator device, and the red arrow indicates the forceps. <a href="https://www.jove.com/files/ftp_upload/65737/65737fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
About 90% of the T. serrulatus scorpions in the Arthropods Bioterium originate from different sites, collected by municipal health agents as part of a surveillance and control program for these animals. The scorpions are caught in urban areas and then sent to Instituto Butantan as part of a collaborative program. T. serrulatus scorpions are parthenogenetic, meaning that birth occurs as a consequence of the development of offspring from unfertilized eggs, without the presence of an individual of the opposite sex12. Due to this type of reproduction, some animals are born in captivity in our facility and are maintained until they reach the adult phase, following a similar husbandry protocol as described below. Once they reach the adult stage, they are added to the venom extraction routine. Upon arrival, the scorpions undergo initial screening, and those in good health conditions are kept in collective enclosures containing a maximum of 350 individuals, divided by their origin. For each enclosure, a control form is filled out daily, providing data on the date of feeding and the type of prey offered (cricket or cockroach), water supply, cleaning procedures, the number of dead animals, and the number of remaining live ones inside the enclosure.
In a pre-scheduled agenda, animals are subjected to electric stimulation, and the resulting venom of hundreds of individuals is then lyophilized, resulting in batches of freeze-dried venom. The standardized venom obtained from this process is destined for horse immunization as part of the manufacturing process, as well as to provide reference material for quality control of the active substance and the final product.
There are very few facilities in the world capable of keeping such a large number of scorpions and performing the volume of venom extractions in accordance with good manufacturing practices and ethical use of the animals, as done in the Arthropods Bioterium of Instituto Butantan. Therefore, our objective is to describe the scorpion maintenance protocols and venom extraction procedures used in T. serrulatus husbandry, which successfully provide the necessary amount of venom for the production of scorpion antivenom.

作者聚焦：优化蝎子毒液提取以生产抗蛇毒血清 

用于抗蛇毒血清生产的 Tityus serrulatus （巴西黄蝎子）的圈养维持和毒液提取

This protocol describes the method to maintain scorpions for the extraction of high amounts of venom, which is used in the production of neutralizing equine-delivered immunoglobulins or antivenoms. There is a high demand for scorpion antivenom in tropical countries, which is the keystone for treating human envenomation cases. The maintainancy procedures reproduce optimal natural conditions, keeping the scorpions in captivity, thus with considering animal welfare and good laboratory practice.In parallel, venom extraction is based on electrical stimulation in the scorpion telson, with minimal harmful effects on the animals. This protocol enabled us to increase exponentially the number of animals maintained in captivity, ranging from 4, 000 to 20, 000 individuals in 10 years. At the same time, venom has been extracted sufficiently to immunize horses to obtain plasma with the specific antibodies, which is the starting material for the manufacturing process that results in around 100, 000 vials of anti-venom per year.The venom milking procedures different from others described in the literature in which the telson is removed, thus limiting the number of extraction per animal. The accumulated experience of the technicians of our facility allow us to extract venom from 200 to 300 animals in around 1 to 1 1/2 hours without anesthetizing the animals or making other invasive procedures.

每个程序都制定了30个标准操作程序（SOP），确保了技术人员之间程序的可重复性并符合质量参数。提取程序后的平均死亡率约为 12%，考虑到电刺激程序和圈养环境，通过应用程序成功降低了压力因素，这可以被认为是低的。
为了保持动物的健康，在盒子的一端放置一个覆盖着棉花和水的托盘，而纸板和蛋托则在另一侧分层放置，模拟 了 T. serrulatus 的自然黑暗和潮湿?...

Watch this Scientific Journal Video about Optimizing Scorpion Venom Extraction for Antivenom Production at JoVE.com

在这里，我们介绍了1922年大规模 的Tityus serrulatus Lutz和Mello（巴西黄蝎）饲养的成功维护和毒液提取方案，目的是为随后的蝎子抗蛇毒血清生产提供毒液，以满足巴西卫生系统的需求。

Scorpion envenomation is a public health problem in several tropical and subtropical countries. Tityus serrulatus Lutz and Mello, 1922 (Brazilian yellow ...

该方案描述了维持蝎子以提取大量毒液的方法，其用于生产中和马递送的免疫球蛋白或抗蛇毒血清。热带国家对蝎子抗蛇毒血清的需求量很大，蝎子抗蛇毒血清是治疗人类毒液中毒病例的基石。维护程序再现了最佳的自然条件，将蝎子圈养起来，从而考虑了动物福利和良好的实验室规范。同时，毒液提取基于蝎子的电刺激，对动物的有害影响最小。该协议使我们能够在10年内成倍地增加圈养动物的数量，从4， 000到20， 000只不等。同时，毒液已被充分提取以对马进行免疫，以获得具有特异性抗体的血浆，这是制造过程的起始材料，每年产生约 100， 000 瓶抗蛇毒血清。毒液挤奶程序与文献中描述的其他程序不同，其中去除了 telson，从而限制了每只动物的提取次数。我们设施技术人员积累的经验使我们能够在大约 1 到 1 1/2 小时内从 200 到 300 只动物中提取毒液，而无需麻醉动物或进行其他侵入性手术。

captive-maintenance-venom-extraction-tityus-serrulatus-brazilian

Research

JoVE Journal

Biology

Captive Maintenance and Venom Extraction of Tityus serrulatus (Brazilian Yellow Scorpion) for Antivenom Production

1.5K 次观看.  Instituto Butantan. 该方案描述了维持蝎子以提取大量毒液的方法，其用于生产中和马递送的免疫球蛋白或抗蛇毒血清。热带国家对蝎子抗蛇毒血清的需求量很大，蝎子抗蛇毒血清是治疗人类毒液中毒病例的基石。维护程序再现了最佳的自然条件，将蝎子圈养起来，从而考虑了动物福利和良好的实验室规范。同时，毒液提取基于蝎子的电刺激，对动物的有害影响最小。该协议使我们能够在1...

视频: 作者聚焦：优化蝎子毒液提取以生产抗蛇毒血清 

Scorpion envenomation is a public health problem in several tropical and subtropical countries. Tityus serrulatus Lutz and Mello, 1922 (Brazilian yellow scorpion) are responsible for approximately 150,000 envenoming cases per year in Brazil, of which 10% require antivenom treatment to reverse life-threatening venom effects. Therefore, thousands of T. serrulatus individuals are maintained under controlled captivity conditions for venom extraction, subsequently used in the production of the national supply of scorpion antivenom. Instituto Butantan is the main antivenom-manufacturing laboratory in Brazil, providing about 70,000 vials of scorpion antivenom for the Brazilian health system. Thus, the husbandry protocols and venom extraction methodologies are key points for the success of large-scale, standardized venom production. The objective of this article is to describe the captivity protocols of T. serrulatus husbandry, encompassing the husbandry routine and the venom extraction procedures, following good manufacturing practices, and ensuring animal welfare. These practices allow for the maintenance of up to 20,000 animals in captivity, with a routine of 3,000 to 5,000 scorpions milked monthly according to antivenom manufacturing demand, achieving an average of 90% of positive extraction.

Here, we present successful maintenance and venom extraction protocols for large-scale Tityus serrulatus Lutz and Mello, 1922 (Brazilian yellow scorpion) husbandry, with the aim of providing venom for subsequent scorpion antivenom production to meet the demand of the Brazilian health system.

科研

生物学

Housing, Captive Maintenance, and Feeding of Brazilian Yellow Scorpions for Venom Extraction

Begin by preparing the enclosures for housing the scorpions. To smoothen the walls of the polypropylene container acting as an enclosure, place 10 centimeters of self adhesive plastic tape over its internal surface, as well as on the upper corners of the container. Then, line the floor of the polypropylene container with craft paper fixed with masking tape on the edges on all sides.Arrange four cardboard trays as vertical substrates on one side of the container with a rigid cardboard sheet between each tray. Place a shallow polypropylene tray containing soaked cotton as a water source on the opposite side of the enclosure. Place the scorpions in the prepared enclosures and house the containers in the room maintained at 24 degrees Celsius.For complete maintenance, first remove the scorpions from the water tray. Then, using long forceps remove the water tray from the enclosure. Place the scorpions in the clean enclosure.Remove all the scorpions left on the enclosure's floor and place them in the clean enclosure. Then remove the remaining live prey from the enclosure. Remove all the disposable substrates from the enclosure and properly discard them before sanitizing the reusable items such as water trays and containers with 0.5%hypochlorite solution.To begin, place the scorpions in communal polypropylene containers as enclosures. Calculate the amount of feed according to the energy intake of the scorpions. Then, count the number of cockroaches as prey items.Then, apply calcium carbonate with alcohol around the top of the enclosure to prevent cockroaches from escaping the enclosure. Place the cockroaches inside the enclosure for two days.

巴西黄蝎的饲养、圈养维护和喂养以提取毒液

Venom Extraction from Brazilian Yellow Scorpions

On the day of venom extraction, count the live scorpions in the enclosure and using long forceps, transfer them to a glass container. Next, arrange all necessary equipment required for venom extraction on the surface of the airflow cabinet. Plug the electro stimulator power cord into the outlet.Then turn on the airflow cabinet, and electro stimulator. Set the displayed parameters on the electro stimulator machine for venom extraction. Using forceps, hold the metasoma of the scorpion, then immobilize the telson with curved forceps before restraining it with a hand.After restraint, place the telson in direct contact with an electro stimulator conductor and collect the venom drops into a plastic micro tube. Transfer the telson from the hand to the curved forceps and gently place the scorpion on the bottom of the glass container. After all the scorpions are milked, place them back into the enclosure.Store the venom at minus 20 degrees Celsius until lyophilization. The venom production of Tityus serrulatus surpasses 80 grams of liquid venom annually equating to over 13 grams of freeze dried venom. Production indicators such as average venom milked per animal, venom extracted per month, and scorpions extracted per month have shown a positive trend line indicating robust production.