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View additional product information for LIVE/DEAD™ Fixable Violet Dead Cell Stain Kit, for 405 nm excitation - FAQs (L34955, L34964, L34963)
18 product FAQs found
有两种简单的方法。一种是通过60°C放置20分钟热灭活细胞。第二种是将细胞放入70%乙醇。乙醇固定的细胞可以在冰箱中无限期储存直到使用，可达好几年。
1.离心细胞、使细胞沉淀并去除上清
2.固定细胞：在15mL含有细胞团的试管中加入10 mL70%冰乙醇，先开始逐滴加入后震荡，混合均匀。
3.在冰箱中储存直到使用。
4.快使用时，水洗两次且在选择的缓冲液中重悬。
•用于流式细胞仪分析的LIVE/DEAD可固定试剂盒适用于固定。这些试剂盒使用胺基反应细胞-非渗透染料对活细胞细胞表面和死细胞胞质染色——活细胞暗淡死细胞明亮。因为染料共价结合到细胞，固定后保留。不幸的是，此方法对基于成像的实验效果不好，由于所有细胞都被染色，很难通过显微镜从暗淡的活细胞中分辨出明亮的死细胞。
•Image-IT DEAD绿色活性染料（货号I10291），用于成像和高通量扫描（HCS）分析的，是一种活细胞非透过型DNA结合染料，固定且透过后可保留长达48小时。
•我们同时推出用于成像分析的LIVE/DEAD Reduced Biohazard Cell Viability试剂盒（货号L7013），其含有两种DNA结合染料，SYTO 10和Dead Red，可以有效染色且通过4%戊二醛固定后快速分析。
注：总的来说，DNA结合的染料和钙黄绿素AM不适用于固定，由于这些染料不能共价地结合到细胞组分上，因此会在固定后缓慢扩散出细胞，逐渐地将所有细胞都染色。
许多抗体和染色剂会标记死细胞。如果您不把死细胞从分析中排除，相关数据将受到干扰。当然，如果你标记的是固定细胞，由于其本身已经死亡，就不再需要添加活性染色剂。但是，如果你们在固定前标记细胞，就需要使用LIVE/DEAD 固定死细胞染色剂。
Attune NxT声波聚焦流式细胞仪检测的下限是0.5 µm。
Attune NxT自动进样器是Attune NxT声波聚焦流式细胞仪的可选附件，能够快速分析至多384份样品。对于不同形式平板——无论96孔还是384孔板——它都具有广泛的兼容性。内置智能探针，最大限度避免堵塞和截留（<0.5%），防止损坏仪器。该仪器通过通气而不是振荡混合，确保样品的均质性，同时维持细胞的活性。它还可作为Attune NxT流式细胞仪关机程序的一个步骤来执行自动清洗。无论采用何种取样方法（试管或平板）和数据采集速率，它都能提供一致的数据。
•模块化设计—多种配置可选，可现场升级。
•节省时间—在不降低数据质量的前提下，速度提高10倍以上。
•简化样品准备流程—无需洗涤，无裂解选项，流体不堵塞。
•可实现独特的应用—可针对广泛的细胞类型和样本构建复杂的实验方案。
针对多参数分析应用，Attune NxT声波聚焦流式细胞仪可以选择最多4个激光器和14色配置，采用模块化系统设计来满足大部分试验需求和实验室预算。创新的光路设计，有助于确保四个空间分隔的激光器精确对齐样品流，确保长期的数据一致性、优越的性能和出色的可靠性。仪器可配备多达4个具有平顶光束的固态激光器（405 nm、488 nm、561 nm和637 nm）。
Attune NxT声波聚焦流式细胞仪的声波聚焦技术采用超声波辐射压力（>2 MHz），将微粒运送至样品流中心。之后，这股预聚焦流注入到为样品提供额外流体动力学压力的鞘液流中。这种“声波辅助流体动力学聚焦”技术可以在任何进样速率的情况下都保证样品流中心范围集中、激光照度均一。在传统的依赖于流体动力学聚焦技术的流式细胞仪中，样品中心需要变宽来适应流速的增加，从而导致激光照度不均一。
细胞计数是一种测量细胞和微粒物理化学特性的方法。流式细胞术可在流式细胞仪内使细胞或微粒单独通过激光来测定其特性，它可用于测定单细胞，针对样品中的每个细胞提供独立的测定结果。此外，还可提供这些样品特性的统计分布结果。 流式细胞仪由三个子系统组成：流体系统、光学系统和电子系统。流体系统负责使细胞移动，从而对其进行分析。光学系统负责产生和收集光信号。电子系统负责将合适的光学信号转化为电子信号，以便进行计算机分析。
Heat killing is commonly used. Place your cells in a tube in buffer and heat at 60oC for 20 minutes. You can also kill your cells by fixing them with ice cold 70% ethanol for 15 minutes. The ethanol-killed cells can then be stored at -20oC until needed, at which point you wash out the ethanol and replace with buffer.
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This dye gives a dim surface label for live cells, but is internalized and gives a brighter signal for dead cells. Flow cytometry is a very sensitive technique and can easily distinguish between the two populations. Microscopy is not as sensitive and may not be able to distinguish the cells because of a less sensitive detector.
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There are two easy options. One is to heat-inactivate the cells by placing at 60 degrees C for 20 minutes. The second is to subject the cells to 70% ethanol. Alcohol-fixed cells can be stored indefinitely in the freezer until use, potentially up to several years.
Centrifuge cells, pellet, and remove supernatant.
Fix cells: Add 10 mL ice cold 70% ETOH to a 15 mL tube containing the cell pellet, adding dropwise at first while vortexing, mix well.
Store in freezer until use.
When ready to use, wash twice and resuspend in buffer of choice.
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The LIVE/DEAD Fixable kits for flow cytometry analysis are compatible with fixation. These kits use amine-reactive cell-impermeant dyes that stain the cell surface of live cells and also the cytosol of dead cells-live cells are dim and dead cells are bright. Since the dye is covalently bound to the cells, it will be retained after fixation. Unfortunately, this method does not work well for imaging-based assays, as all cells are stained and it is difficult to distinguish bright dead cells from dim live cells with a microscope. Ethidium monoazide (EMA; Cat No. E1374) is a cell impermeant nucleic acid stain that can be applied to live cultures and stains only dead cells. After incubation and washing away unbound dye, the cells can be exposed to light to photoactivate EMA to crosslink to dead cell DNA. After crosslinking to dead cell DNA, the samples may be fixed and permeabilized.
Image-IT DEAD Green Viability Stain (Cat. No. I10291) for imaging and high-content screening (HCS) analysis is a live-cell impermeant DNA binding dye that is compatible with fixation and permeabilization with good retention up to 48 hours.
We also have a LIVE/DEAD Reduced Biohazard Cell Viability Kit (Cat. No. L7013) for imaging and flow analysis that contains two DNA binding dyes, SYTO 10 and Dead Red, that are sufficiently retained to be analyzed soon after 4% glutaraldehyde fixation.
Note: In general, DNA-binding dyes and calcein AM are not compatible with fixation, as these dyes are not covalently bound to components of the cell and will thus slowly diffuse out of cells after fixation, gradually staining all cells as dead.
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Many antibodies and stains will label dead cells. This will give you misleading data if you do not exclude the dead cells from your analysis. Of course, if you are labeling fixed cells, they are already dead and you do not need a viability stain. However, if you label your cells prior to fixation, then you need to use one of the LIVE/DEAD Fixable Dead Cell Stains.
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The smallest size that you can detect with the Attune NxT Acoustic Focusing Cytometer is 0.5 µm.
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The Attune NxT Autosampler, an optional accessory for the Attune NxT Acoustic Focusing Cytometer, enables rapid processing of up to 384 samples. It has broad compatibility with different plate formats, both 96- and 384-well plates. It has an intelligent probe designed to minimize clogging and carryover (<0.5%) and to prevent damage to the instrument. It mixes by aspiration rather than shaking to ensure homogeneity of the sample and maintain cell viability. Is performs automated cleaning as part of the shutdown process of the Attune NxT Cytometer. It provides consistent data regardless of sampling method (tube vs. plate) and collection rate.
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-Modular design - Multiple configurations available - field upgradable.
-Save time - 10X faster speeds with no loss in data quality.
-Simplified sample prep - No wash, no lyse options, non-clogging fluidics.
-Enables unique applications - Complex protocols on a broad range of cell types and samples.
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With the option to be configured with up to 4 lasers and 14 colors for multi-parameter analysis the Attune NxT Acoustic Focusing Cytometer was designed as a modular system to fit most experimental needs and lab budgets. The novel design of the optical path helps ensure precise fixed alignment of four spatially separated lasers onto the sample stream enabling consistency in data over time, superior performance, and superior reliability. The instrument can be configured with up to 4 solid-state lasers (405 nm, 488 nm, 561 nm, and 637 nm) with flat top beam profiles.
The Attune NxT Flow Cytometer's acoustic focusing uses ultrasonic radiation pressure (>
2 MHz) to transport particles into the center of the sample stream. This pre-focused stream is then injected into the sheath stream, which supplies an additional hydrodynamic pressure to the sample. The combination of these two forces- termed acoustic-assisted hydrodynamic focusing-results in a narrow core stream and uniform laser illumination, regardless of the sample input rate. In traditional cytometers that rely solely on hydrodynamic focusing, the sample core widens to accommodate the increases in flow rate, which results in less uniform laser light illumination.
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Cytometry is the measurement of physical or chemical characteristics of cells or particles. Flow cytometry measures these characteristics of cells or particles as they individually pass lasers in a flow cytometer instrument. Flow cytometry is performed on single cells, providing discrete measurements for each cell in the sample. It also provides a statistical distribution of the measured characteristics of the sample.
A flow cytometer is made up of three subsystems: fluidics, optics, and electronics. Fluidics moves the cells and introduces them for interrogation. Optics generates and collects the light signals. Electronics converts the optical signals to proportional electronic signals for computer analysis.
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