NIR 蛋白质印迹 NewBlot™ IR 剥离缓冲液,NewBlot IR 去除所有 IRDye 二抗

Reagents > NewBlot™ IR Stripping Buffers for NIR Western Blots,用于 NIR 蛋白质印迹的 NewBlot™ IR 剥离缓冲液

NewBlot™ IR Stripping Buffers for NIR Western Blots

NewBlot IR is a robust formulation optimized for near-infrared (NIR) Western blots that works with both nitrocellulose and PVDF membranes. This stripping buffer is a good place to start if you are new to stripping and reprobing NIR Western blots.

NewBlot IR works well for removal of all IRDye secondary antibodies, and may be suitable for removal of other dye-conjugated secondary antibodies.

NewBlot IR does not require hazardous shipping, unlike many other stripping buffers.

Note: NewBlot IR Stripping Buffer is not recommended for loading amounts over 30 µg.

Choosing the Right Stripping Buffer

NewBlot IR 是一种针对近红外 (NIR) 蛋白质印迹优化的强大配方，适用于硝酸纤维素膜和 PVDF 膜。 如果您不熟悉剥离和重新检测 NIR 蛋白质印迹，则此剥离缓冲液是一个很好的起点。

NewBlot IR 适用于去除所有 IRDye 二抗，并且可能适用于去除其他染料偶联二抗。

与许多其他剥离缓冲液不同，NewBlot IR 不需要危险运输。

注意：不建议将 NewBlot IR 剥离缓冲液用于上样量超过 30 µg。

选择正确的剥离缓冲液

Specifications

NewBlot IR Stripping Buffer is supplied as a 5X concentrated solution. Each 100 mL bottle is sufficient for up to 3000 cm2 or approximately fifty 7 × 8.5 cm Odyssey® nitrocellulose membranes.

NewBlot IR 剥离缓冲液以 5X 浓缩溶液的形式提供。 每个 100 mL 瓶足以容纳高达 3000 cm2 或大约 50 个 7 × 8.5 cm Odyssey® 硝酸纤维素膜。

Example Data

NewBlot™ IR Stripping Buffers for NIR Western Blots
Stripping and reprobing nitrocellulose or PVDF membranes effectively with NewBlot IR Stripping Buffer. EGFR and phospho-ERK levels were compared in EGF-stimulated (+) and non-stimulated (-) A431 lysate (1 µg total protein). The membrane was probed with mouse anti-EGFR, mouse anti-pERK1/2, and Β-tubulin rabbit polyclonal (P/N 926-42211), then detected with IRDye 800CW Goat anti-Mouse (P/N 926-32210) and IRDye 680RD Goat anti-Rabbit (P/N 926-68071). The blot was scanned with an Odyssey CLx Imaging System (Original blot). The blot was then stripped with NewBlot IR Stripping Buffer (P/N 928-40028), and scanned again (Strip #1). The blot was detected with the same primary and secondary antibodies and scanned again (Reprobe #1). The process was repeated 2 more times (Strip and Reprobe #2-3). Image display settings for all stripped and reprobed images are identical to the original image.
使用 NewBlot IR 剥离缓冲液有效剥离和重新检测硝酸纤维素或 PVDF 膜。 比较了 EGF 刺激 (+) 和非刺激 (-) A431 裂解物（1 µg 总蛋白）中的 EGFR 和磷酸化 ERK 水平。 用小鼠抗 EGFR、小鼠抗 pERK1/2 和 β-微管蛋白兔多克隆 (P/N 926-42211) 探测膜，然后用 IRDye 800CW 山羊抗小鼠 (P/N 926-32210) 和 IRDye 680RD 山羊抗兔 (P/N 926-68071)。 使用 Odyssey CLx 成像系统（原始印迹）扫描印迹。 然后用 NewBlot IR 剥离缓冲液 (P/N 928-40028) 剥离印迹，并再次扫描 (Strip #1)。 用相同的一抗和二抗检测印迹并再次扫描（重新探针 #1）。 该过程再重复 2 次（Strip and Rerobe #2-3）。 所有剥离和重新探测图像的图像显示设置与原始图像相同。

Factors Affecting Efficiency when using NewBlot IR Stripping Buffer

Recommended Standard Conditions: 1X NewBlot IR for 20 minutes at room temperature.

Below are key factors that affect stripping efficiency with NewBlot IR on Odyssey nitrocellulose or Immobilon®-FL PVDF membranes.

For optimal stripping results, follow the optimization guidelines in NewBlot IR Stripping Buffer pack insert.

Amount of time blot is in stripping buffer

Increasing the stripping time may increase stripping efficiency. However, it may also remove target proteins and reduce the success of reprobing.

推荐的标准条件：1X NewBlot IR 在室温下 20 分钟。

以下是影响使用 NewBlot IR 在 Odyssey 硝酸纤维素膜或 Immobilon®-FL PVDF 膜上剥离效率的关键因素。

为获得最佳剥离结果，请遵循 NewBlot IR 剥离缓冲液包插入中的优化指南。

印迹在剥离缓冲液中的时间量
增加汽提时间可以提高汽提效率。 然而，它也可能去除目标蛋白并降低重新探测的成功率。

Sample type and preparation

Even under the most stringent stripping conditions, the fluorescent signal may not be removed completely due to sample load amount, antibody affinity/avidity, and target protein abundance.

Note: NewBlot IR Stripping Buffer is not recommended for loading amounts over 30 µg.

即使在最严格的剥离条件下，由于样品上样量、抗体亲和力/亲合力和靶蛋白丰度，荧光信号也可能无法完全去除。

注意：不建议将 NewBlot IR 剥离缓冲液用于上样量超过 30 µg。

Blot handling conditions

Washing, scanning, or stripping efficiency will be affected if the blot is allowed to dry at all during incubation. Keep the blot moist at all times.

Buffer concentration and temperature used for stripping

Increasing the stripping buffer concentration or temperature will reduce stripping efficiency and is not recommended for NewBlot IR Stripping Buffer. Use the material at a 1X concentration at room temperature for optimal results.

印迹处理条件
如果在孵育过程中让印迹完全干燥，洗涤、扫描或剥离效率将受到影响。 始终保持印迹湿润。

用于剥离的缓冲液浓度和温度
增加剥离缓冲液浓度或温度会降低剥离效率，不推荐用于 NewBlot IR 剥离缓冲液。 在室温下使用 1X 浓度的材料以获得最佳结果。

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