NEW YORK – Countable Labs, formerly known as Enumerix, expects its chemistries and benchtop reader will allow researchers to measure single DNA molecules quickly and cheaply.

纽约——Countable Labs（前身为Enumerix）预计其化学技术和台式读取器将使研究人员能够快速且廉价地测量单个DNA分子。

After recently rebranding, the Palo Alto, California-based startup will launch its first product, called Countable PCR, at the American Society of Gene & Cell Therapy meeting next week.

最近更名的加州帕洛阿尔托的这家初创公司，将在下周的美国基因与细胞治疗学会会议上推出其首款产品——Countable PCR。

The firm has developed a

公司已经开发了一项

method

方法

to create 30 million tiny compartments within a gel matrix in a standard Eppendorf tubestrip, all within about 20 minutes.

在标准的Eppendorf管条中，在约20分钟内在凝胶基质中创建3000万个微小隔间。

When applied to PCR reactions, the approach is similar to digital PCR but obviates the need for standard curves or Poisson statistics.

应用于PCR反应时，该方法类似于数字PCR，但无需标准曲线或泊松统计。

Although the firm's first application for its technology is PCR, Countable plans to expand into other assays. 'We consider our technology essentially like a picoliter bioreactor,' said Jocelyn Davé, the firm's new CEO. 'The simplest way to measure something is counting, and that's what we're doing.'.

尽管该公司技术的首个应用是PCR，Countable计划扩展到其他检测领域。公司新任首席执行官乔斯琳·达维表示：“我们的技术本质上就像一个皮升生物反应器。测量事物最简单的方法就是计数，而这正是我们在做的。”

Eleen Shum, Countable's founding scientist and VP of product development, said the Countable approach, along with its simple workflow and automated results, can potentially speed up scientific projects, at a lower cost than standard methods.

Countable的创始科学家兼产品开发副总裁许伊琳表示，Countable的方法结合其简单的工作流程和自动化的结果，有可能以比标准方法更低的成本加快科学项目的速度。

For example, the 30 million reaction compartments allow researchers to do their work using fewer total reactions, she said. The approach also yields error bars with a CV of approximately 1 percent, a 10-fold improvement over digital PCR.

她说，例如，3000万个反应隔室可以让研究人员用更少的总反应来进行他们的工作。该方法还产生了大约1%的变异系数的误差条，比数字PCR提高了10倍。

The approach has no dead volume or sample loss, as it uses a centrifugation-based method rather than microfluidics. The gel block can also be stored in the fridge and re-imaged, and the firm is currently perfecting a technique with a collaborator that will potentially enable users to harvest the contents of each microreactor, removing and reuniting the whole sample from the gel block.

该方法没有死体积或样品损失，因为它使用的是基于离心的方法，而不是微流体。凝胶块也可以存放在冰箱中并重新成像，该公司目前正在与合作伙伴完善一项技术，这项技术有望让用户能够提取每个微反应器的内容物，将整个样品从凝胶块中取出并重新组合。

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Countable has also developed a novel multiplexing chemistry called Universal Multiplexing, which Davé said is 10 times less expensive than standard approaches. The technique was described in a

可数公司还开发了一种名为通用多重化的新型多重化学技术，Davé表示该技术的成本比标准方法低10倍。该技术在一份报告中有所描述。

BioRxiv

生物预印本

publication two years ago, along with a proof-of-concept panel enabling simultaneous detection of 22 pathogen targets.

两年前发表，同时提供了一个可同时检测22个病原体目标的概念验证面板。

Combining Universal Multiplexing with Countable PCR's dynamic range of up to 1 billion molecules could also potentially shorten project times for diagnostics developers  by allowing them to detect both rare and abundant targets in a single reaction with minimal optimization.

结合通用多重技术和可计数PCR高达10亿分子的动态范围，还可以通过允许诊断开发者在单次反应中同时检测稀有和丰富目标、且几乎无需优化，从而潜在缩短项目时间。

In addition to the proprietary picoliter reaction vessels, the method uses its own consumables and reagents, including a spin column and PCR mix, with the PCR reaction processed on any standard thermal cycler.

除了专有的皮升反应容器外，该方法还使用其自身的耗材和试剂，包括离心柱和PCR混合液，并在任何标准的热循环仪上进行PCR反应处理。

Readout is done on the firm's own instrument, which can process between one and 96 samples, with four color channels. It relies on a 3D scanning technique called light sheet microscopy, which essentially performs optical z-sectioning of a sample.

读取是在公司自己的仪器上完成的，该仪器可以处理1到96个样本，并具有四个颜色通道。它依赖于一种称为光片显微镜的3D扫描技术，该技术本质上对样本进行光学Z轴切片。

In a poster at ASGCT, the Countable team also highlighted ways its tech may be more versatile and approachable compared to legacy methods for rare molecule detection in a high background ─ specifically for viral titer characterization.

在ASGCT的一张海报中，Countable团队还强调了其技术相较于传统方法在高背景下的罕见分子检测方面可能更加通用且易于操作——特别是针对病毒滴度的表征。

For this application, qPCR may be too variable, while dPCR can suffer from a narrow dynamic range, high Poisson errors, and arbitrary thresholding, the firm said. In addition, conducting both viral titer and genome integrity assessments in a single assay is challenging using dPCR due to a relatively smaller number of partitions or droplets..

该公司表示，对于这一应用，qPCR的变异性可能太大，而dPCR则可能受到动态范围窄、泊松误差高和阈值设定任意的困扰。此外，由于分区或液滴的数量相对较少，使用dPCR在单一检测中同时进行病毒滴度和基因组完整性评估具有挑战性。

In contrast, in its ASGCT poster, Countable demonstrated a novel multiplex assay that interrogates different regions of a gene of interest to quantify the ratio of partial and complete transgenes while simultaneously measuring viral titer.

相比之下，在其ASGCT海报中，Countable展示了一种新型的多重检测方法，该方法通过检测目标基因的不同区域来量化部分和完整转基因的比例，同时测量病毒滴度。

Unlike dPCR, the single-molecule counting approach 'prevents false co-occupancy of fragmented, unlinked targets, ensuring accurate genome integrity data,' according to the poster. Full genome integration can be further validated with long-read amplicon analysis, as the firm demonstrated up to 2,000 bp length.

根据海报，与dPCR不同，单分子计数方法“防止了片段化、未连接目标的假性共定位，确保准确的基因组完整性数据”。全基因组整合可以通过长读长扩增子分析进一步验证，该公司已展示长达2000个碱基对的长度。

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The Countable team also presented two posters at the Association for Molecular Pathology meeting last November.

去年十一月，Countable 团队还在分子病理学协会会议上展示了两份海报。

The first demonstrated the technologies' capability to 'expand the haystack,' Shum said, by counting molecules from 3 μg of DNA. This method could be particularly useful for assays using tissue or whole blood samples.

舒姆表示，第一个演示展示了这项技术“扩展样本规模”的能力，通过计算来自3微克DNA的分子数量。这种方法对于使用组织或全血样本的检测可能特别有用。

The second poster showed how Countable PCR could increase confidence in rare molecule detection, which the firm suggested makes it ideal for cell-free DNA, rare cells in cerebrospinal fluid, or other low-input samples.

第二张海报展示了可计数PCR如何提高对稀有分子检测的信心，该公司表示，这使其非常适合用于无细胞DNA、脑脊液中的稀有细胞或其他低输入样本。

First user insights

初步用户洞察

Rick Tarleton, a cell biologist at the University of Georgia, learned about Countable Labs from its proof-of-concept study in

乔治亚大学的细胞生物学家里克·塔尔顿从其实验室的概念验证研究中了解到了Countable Labs。

Analytical Chemistry

分析化学

. In his efforts to develop a diagnostic for Chagas disease, he had been searching for a way to perform many PCR assays as quickly and cheaply as possible.

在他的努力开发查加斯病诊断方法的过程中，他一直在寻找一种尽可能快速和廉价地进行大量PCR检测的方法。

Chagas disease is caused by the protozoon

查加斯病是由原生动物引起的

Trypanosoma cruzi

克鲁斯锥虫

. The parasite is transmitted in saliva from the bite of a reduviid bug — also known as the kissing bug because it commonly bites the face — and it is most common in rural, impoverished areas of South America. Chronic

这种寄生虫通过一种叫做锥蝽（也被称为接吻虫，因为它通常叮咬面部）的虫子唾液传播，最常见的传播地区是南美洲农村贫困地区。慢性

T. cruzi

克鲁兹锥虫

infections involving the heart and digestive system can be ultimately fatal.

涉及心脏和消化系统的感染最终可能致命。

Tarleton and his team at the Center for Tropical and Emerging Global Diseases are now using Countable PCR for ultrasensitive detection and quantification of

塔勒顿和他的团队在热带和新兴全球疾病中心现在正在使用可计数PCR进行超灵敏检测和定量

T. cruzi

克鲁兹锥虫

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In a recent

最近在

publication

出版物

, they described a method to perform several hundred standard PCR detection reactions to increase low-level detection of the parasite in host insects'This approach works well, but is laborious and expensive,' Tarleton said, so the team has now been working with Countable Labs instead.

，他们描述了一种进行数百次标准PCR检测反应的方法，以提高在宿主昆虫中对寄生虫的低水平检测。这种方法效果很好，但既费力又昂贵，”塔勒顿说，因此该团队现在已转而与Countable Labs合作。

According to Tarleton, the major advantages of the Countable method are the high DNA loading per reaction compared to standard PCR and the wide dynamic range. 'For hosts with this infection, there is a greater than millionfold difference in parasite load,' he said, but the Countable Labs method 'covers this range with a single DNA input.' .

根据塔尔顿的说法，Countable 方法的主要优势在于每次反应的高 DNA 装载量，相比标准 PCR 具有更宽的动态范围。“对于感染这种病原体的宿主，寄生虫载量差异超过百万倍，”他说道，但 Countable Labs 的方法“只需单次 DNA 输入即可覆盖这一范围。”

The team is also impressed with the multiplexing capability and hopes that using multiple targets will improve sensitivity, as well, Tarleton said.

Tarleton 表示，该团队对多路复用能力也印象深刻，并希望使用多个目标也能提高灵敏度。

Although they've been focused on the Chagas disease application, Tarleton expects to have other experimental applications for the Countable system in the future, such as detection of

尽管他们一直专注于查加斯病的应用，但塔勒顿预计未来 Countable 系统还会有其他实验性应用，例如检测。

T. cruzi

克氏锥虫

in animals and humans. They are hoping to ultimately establish a clinical diagnostic lab that can offer the test for other research studies. 'This is an extremely important need that is not being met by any other technology,' he said.

在动物和人类身上。他们希望最终建立一个临床诊断实验室，可以为其他研究提供这种测试。他说：“这是一个非常重要且未被任何其他技术满足的需求。”

Regarding federal funding cuts impacting global public health research, Tarleton said his team was fortunate to get in under the wire with new NIH funding for developing an assay on the Countable Labs system, allowing them to continue the project for the time being. 'Unfortunately, the recent changes in policy involving foreign [collaborators] on NIH grants is going to make it difficult to demonstrate the broad utility of this approach for human infections,' he said, since they are more common in Central and South America.

关于联邦资金削减对全球公共卫生研究的影响，塔雷顿表示他的团队很幸运能够赶在最后一刻获得美国国立卫生研究院（NIH）的新一轮资助，用于在Countable Labs系统上开发检测方法，使他们目前能够继续该项目。 “不幸的是，最近涉及NIH拨款中外国[合作者]的政策变化，将使得证明这种方法对于人类感染的广泛适用性变得困难。”他说，因为这些感染在中美洲和南美洲更为常见。

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Countable Labs plans to install a new system in Tarleton's lab next month, Shum said, and will continue to collaborate with him to develop a high-sensitivity test for pathogen detection.

舒姆说，Countable Labs 计划下个月在塔勒顿的实验室安装一个新系统，并将继续与他合作开发一种用于病原体检测的高灵敏度测试。

Going forward, Countable aims to also address the oncology research market, as well as clinical applications.

展望未来，Countable 还旨在解决肿瘤学研究市场以及临床应用问题。

'We hope to grow and enable more applications,' Davé said, since the technology's sensitivity 'can enable more testing, earlier testing, and even more routine testing, as well.'

“我们希望发展壮大并实现更多的应用，”戴夫说，因为该技术的敏感性“可以实现更多、更早甚至更常规的测试。”