返回重点引用Fluoro-Gold™ 标记的视网膜外植体培养用于神经毒性筛查研究相关页面细胞染色细胞表面、细胞核、线粒体、溶酶体等的荧光染色分享文章与同事分享这篇文章。复制链接发送电子邮件视网膜和视神经疾病，可导致失明, 由于人口老龄化，患病率正在增加。幸运的是，对这些眼部疾病的快速了解正在产生新的治疗方法和药物输送系统。这就需要改进眼部药物临床前药物毒性筛选方法。在最近一期的氧化医学和细胞寿命中，作者 Smedowski 等。阿尔。描述使用大鼠视网膜外植体培养物的药物筛选系统。在分离外植体之前，视网膜神经节细胞用 Fluoro-Gold™ 进行荧光标记，Fluoro-Gold™ 是一种神经元逆行示踪剂，可促进视网膜神经元的定量评估植物。为了测试该系统，外植体暴露于有毒浓度的庆大霉素，这会导致视网膜细胞损伤并引发乳酸脱氢酶 (LDH) 快速释放到培养基中，并降低视网膜神经节细胞密度。此外，补充睫状神经营养因子（一种已知的神经保护剂）被证明可以最大限度地减少细胞损伤并减少 LDH 的释放。该系统被发现是一种快速、可重复且灵敏的检测视网膜药物筛选研究中神经毒性的方法；以及通过体内玻璃体内药物递送进行筛选的可行替代方法。经过 7 天培养后，FluoroGold 标记的整体器官型视网膜外植体（来自 Smedowski 等人，2018 年）。Biotium 销售超纯 Fluoro-Gold™（也称为Hydroxystilbamidine）。我们还提供多种其他荧光神经末梢和神经元示踪剂。Fluoro-Gold 是 Fl​​uorochrome, LLC 的商标。返回重点引用PMAxx™ q-PCR：一种检测种子传播病原体的有效工具相关页面生存力 PCRA 通过 PCR 选择性和灵敏地定量活细菌的强大方法分享文章与同事分享这篇文章。复制链接发送电子邮件种子健康对于良好的产量和维持所需的植物性状，以及最大限度地减少植物病原体的传播和扩散。无病原体种子认证是预防和管理疾病的有效策略。然而，这种方法高度依赖于可靠的病原体检测方法。Clavibacter michiganensis subsp。 michiganensis (Cmm) 是一种种子传播的病原体，可导致番茄青枯病和溃疡病。这是一种具有严重经济后果的毁灭性疾病。多种技术，包括将实时定量 PCR (qPCR) 与基于培养的分离相结合的生物 PCR，已被用于提高 Cmm 检测的准确性。然而，在 th 中检测 Cmm可行但不可培养的 (VBNC) 状态仍然具有挑战性。 PLOS ONE 最近的一篇论文展示了一种改进且有效的策略，可以检测活体和 VBNC 状态下的 Cmm，而使用传统方法无法检测到 Cmm。该研究证实，与 PMA 相比，新改进的光活化染料 PMAxx™ 具有更高的区分活细胞和死细胞的灵敏度，并且与活细胞 DNA 的结合更少。作者还描述了用于准确 Cmm 检测的 PMAxx™ 的最佳浓度、细菌 CFU/mL、样品制备和光解条件。病原体：Clavibacter michiganensis 亚种。 michiganensis (S. Nelson, Flickr) 了解有关 Viability PCR 技术的更多信息，包括 viability PCR 染料、试剂、试剂盒和光解设备。From: 106Sizes: 5-20 ug labeling, 10-20 ug labeling, 20-50 ug labeling, 25-50 ug labeling, 50-100 ug labelingCatalog #: 92433, 92432, 92431, 92430, 92429, 92428, 92427, 92426, 92425, 92230, 92231, 92232, 92233, 92234, 92235, 92236, 92237, 92238, 92239, 92240, 92241, 92242, 92243, 92245, 92246, 92247, 92248, 92250, 92251, 92252, 92253, 92254, 92255, 92256, 92257, 92258, 92259, 92260, 92261, 92262, 92263, 92264, 92265, 92267, 92268, 92270, 92271, 92272, 92273, 92274, 92275, 92276, 92277, 92278, 92279, 92280, 92281, 92282, 92283, 92284, 92285, 92287, 92288, 92289, 92290, 92291, 92303, 92304, 92305, 92316, 92317, 92318, 92319, 92320, 92321, 92322, 92323, 92324, 92331, 92332, 92333, 92334, 92335, 92336, 92337, 92338, - 92339View allHide From: 120Sizes: 5-20 ug labeling, 20-50 ug labelingCatalog #: 92501, 92500, 92502, 92503, 92504, 92505, 92506, 92507, 92508, 92509, 92510, 92511, 92512, 92513, 92514, - 92515View allHide A high-performance dye-based qPCR master mix containing EvaGreen®, premixed ROX, Cheetah HotStart Taq Polymerase, and a low concentration of the Forget-Me-Not inert blue tracking dye. EvaGreen® Dye and Master Mixes What is the difference between EvaGreen® and SYBR® Green?

EvaGreen® dye is spectrally similar to FAM or SYBR® Green I, which means no change in optical settings is required for using an EvaGreen®-based master mix. However, there are several differences between the dyes.

Lower background fluorescence: EvaGreen® dye has less background than SYBR® Green I due to its novel \"release-on-demand” DNA-binding mechanism.

Brighter and non-inhibitory:  EvaGreen® is less inhibitory in the PCR reaction than SYBR® Green, permitting the use of saturating dye concentration for maximal signal and better high-resolution DNA melt analysis.

Safer and more environmentally friendly: EvaGreen® dye is the first and only PCR dye to date designed to be environmentally safe.

Dye stability: EvaGreen® dye is very stable both during storage and under PCR conditions. SYBR® Green I, on the other hand, is known to degrade following multiple freeze-thaw cycles and under PCR conditions. Moreover, decomposed SYBR® Green I is reported to be even more inhibitory to PCR.

Digital PCR: EvaGreen® dye is currently the only qPCR dye to be used in droplet digital PCR (ddPCR).

Category: EvaGreen® Dye and Master MixesView more FAQs How do I read EvaGreen® fluorescence in qPCR?

EvaGreen® dye is spectrally similar to FAM or SYBR® Green I, which means no change in optical settings is required for using an EvaGreen®-based master mix compared to SYBR® Green-based master mixes. You can read your signal in the green fluorescence channel (FAM, SYBR® Green, or EvaGreen®) of your instrument.

Category: EvaGreen® Dye and Master MixesView more FAQs Which EvaGreen® product, 20X in water or 2000X in DMSO, is best for qPCR?

For most qPCR applications, we recommend using EvaGreen® Dye, 20X  in water (31000), or one of our 2X EvaGreen® qPCR Master Mixes. EvaGreen® Dye, 2000X  in DMSO (31019) is offered for specialized applications requiring a concentrated dye solution.

Category: EvaGreen® Dye and Master MixesView more FAQs

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2X Forget-Me-Not EvaGreen® qPCR Master Mix is a ready-to-use high performance dye-based master mix for qPCR and DNA melt curve analysis. The 2X Master Mix contains EvaGreen® dye, premixed ROX, Cheetah HotStart Taq DNA Polymerase, dNTPs, and a low concentration of the Forget-Me-Not inert blue tracking dye.

Ready-to-use, just add primers and DNA templateOptimized with our superior EvaGreen® Dye which is more sensitive than SYBR® Green I and other commercial PCR dyesPrevent wasted samples and other costly mistakes with our Forget-Me-Not light blue master mixPremixed with ROX and optimized for instruments requiring either high or low concentrations of ROX reference dyeFormulated with Cheetah HotStart Taq for fast cycling protocolsDirectly visualize amplification products in a gel

Figure 1: Forget-Me-Not EvaGreen® qPCR Master Mix contains a low concentration of an inert blue dye, which allows the user to visually distinguish wells containing reaction mix from empty wells, and can thereby reduce pipetting errors, saving time and reagents.Forget-Me-Not Color Tracking

Catch pipetting mistakes and avoid wasting time, reagents, and your precious DNA samples. Forget-Me-Not EvaGreen® qPCR Master Mixes with premixed ROX contains a low concentration of the Forget-Me-Not inert blue dye. The dye does not interfere with PCR or signal acquisition but does allow the user to easily distinguish wells containing reaction mix from empty wells (Figure 1). This product is for users who wish to use single color tracking. Users that require 2-color tracking should purchase Forget-Me-Not EvaGreen® qPCR Master Mix (2-Color Tracking) which features the 40X Forget-Me-Not Template Buffer. When the 40X Forget-Me-Not Template Buffer is mixed with your DNA template and added to the reaction, it turns dark blue, allowing you to see at a glance which wells have template added.

Peace of Mind Plus PCR Performance

Forget-Me-Not qPCR Master Mix has been formulated for fast cycling PCR parameters, but can also be used with regular cycling protocols. Forget-Me-Not qPCR Master Mix performs as well as our Fast EvaGreen® Master Mix, and as well or better than Qiagen’s QuantiNova® SYBR® Green PCR Master Mix in a real-time PCR assay (Figure 2), while making it easy for you to make sure your reactions are set up correctly the first time, every time.

Figure 2. Real-time PCR data comparing Forget-Me-Not (blue lines) with Biotium s Fast EvaGreen® (green lines) and Qiagen s QuantiNova® SYBR® Green (gray lines) master mixes. A. Amplification curves on linear scale. EvaGreen® dye-based master mixes yield higher signal compared to the SYBR® Green-based mix. B. Amplification curves on logarithmic scale. Forget-Me-Not performs as well or better than the other master mixes. NTCs: non-template controls (dashed lines).EvaGreen® Dye

An environmentally friendly, non-mutagenic, and non-cytotoxic DNA-binding dye with features ideal for both qPCR and High Resolution Melt® (HRM) analysis (Ref. 3). EvaGreen® Dye binds to dsDNA via a novel \"release-on-demand” mechanism in which the dye is in an inactive conformation until DNA is available. As DNA is produced during the PCR process, the dye shifts to an active conformation that binds DNA to emit fluorescence. As a result, EvaGreen® Dye can be used at a much higher dye concentration in qPCR than SYBR® Green I (Ref. 1). The dye has excitation and emission spectra very close to those of fluorescein (FAM) or SYBR® Green I, making it readily compatible with instruments equipped with the 488 nm argon laser or any visible light excitation with wavelength in the region. EvaGreen® amplification products can be directly detected on a gel without the need for another DNA gel stain.

Independent labs have confirmed that EvaGreen® dye is non-mutagenic, non-cytotoxic, and safe to aquatic life for direct disposal in the drain. Other commercial PCR dyes enter into cells in a matter of minutes. SYBR® Green I, for example, has been shown to be environmentally more toxic than ethidium bromide, a well-known mutagen (Ref. 2). For details, download the EvaGreen® Dye Safety Report.

Learn more about EvaGreen® Dye for qPCR.

Cheetah HotStart Taq

Cheetah HotStart Taq DNA Polymerase is Biotium’s proprietary chemically-modified hotstart DNA Polymerase. Cheetah Taq is completely inactive at room temperature, and is fully activated after 2 minutes at 95°C, making it particularly suitable for fast cycling PCR protocols.

Forget-Me-Not EvaGreen® qPCR Master Mix with Premixed ROX is available in two formulations: a low ROX master mix (~50 nM final ROX concentration in assay) and a high ROX master mix (~500 nM final assay concentration). ROX is required as a passive reference dye in some qPCR systems to compensate for well-to-well optical variations. Refer to the table below for the passive dye requirements of real-time PCR instruments.

Low ROX(~50 nM)Applied Biosystems®: 7500, 7500 Fast, ViiA™7, Quant-Studio™ instrumentsStratagene (Agilent): MX4000P, MX3000P, MX3005P High ROX(~500 nM)Applied Biosystems®: 5700, 7000, 7300, 7700, 7900, 7900HT, 7900HT Fast, StepOne™, StepOnePlus™ No ROX required*BioRad: iCycler™, MyiQ™, MiQ™ 2, iQ™ 5, CFX-96 Touch™, CFX-384 Touch™ and Connect™, Chromo4™, MiniOpticon™Qiagen: Rotor-Gene® Q, Rotor-Gene® 3000, Rotor-Gene®6000Eppendorf: Mastercycler® RealplexIllumina: Eco™ RealTime PCR SystemCepheid: SmartCyler®Roche: LightCycler® 480, LightCycler® 2.0*Instruments that do not require ROX reference dye are generally compatible with qPCR master mixes containing ROX (check with the manufacturer before use). We also sell Forget-Me-Not EvaGreen® qPCR Master Mix kits without ROX (cat# 31041), and with a separate tube of ROX (cat# 31042) that can be used to make your own low or high ROX master mixes.**Bio-Rad’s iCycler users do not need to add fluorescein to the PCR reaction as EvaGreen® dye has a slight background fluorescence that provides adequate and stable baseline level fluorescence. For these instruments we recommend using Forget-Me-Not EvaGreen® qPCR Master Mix without ROX (cat# 31041).

EvaGreen Dye and Cheetah Taq are covered under US and international patents. QuantiNova is a registered trademark of Qiagen Group. SYBR is a registered trademark of Thermo Fisher Scientific. HRM is a registered trademark of Idaho Technologies, Inc./BioFire Defense, LLC; the use of HRM may require a license.

1. BMC Biotechnol (2007) https://doi.org/10.1186/1472-6750-7-762. Mutation Res. (2001) https://doi.org/10.1016/S1383-5718(01)00155-33. Parasites Vectors (2019) https://doi.org/10.1186/s13071-019-3781-4

A high-performance product for fluorescent probe-based PCR applications, including quantitation and SNP genotyping. Detailed ViewAn easy and accurate way to determine the activity of a nucleic acid polymerase without using radioisotopes. Detailed ViewGelGreen® is a sensitive, non-mutagenic and environmentally safer green fluorescent DNA gel stain. Detailed ViewA high-performance dye-based qPCR master mix containing EvaGreen®, Cheetah HotStart Taq Polymerase, and the Forget-Me-Not 2-color tracking system. Detailed ViewSuperior RNA extraction from FFPE samples. 3 to 30-fold increase in yield. 2-fold increase in DV200 (RNA over 200 nuc). Detailed ViewBiotium implements a Quality System, certified by QAS according to Standard QAS ISO 9001:2015. Certificate No. US3564

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