（ HRP-Polymer）免疫组化试剂盒（Rabbit）是高敏感性试剂盒。用聚合标记方法将过氧化物酶连接到二抗上，形成超大分子抗体-酶聚合物，替代传统方法中的二抗和三抗。这种聚合物拥有强大的信号放大能力，同时有很强的组织细胞渗透力，特别适合免疫组化的应用。此系列检测试剂与传统的三步法比较具有简单，快速，高敏感，低背景等特点。
包装清单：
R777876
| Component
| 1 Kit
| Storage
|
| R777876A | 封闭液（5%BSA） | 6mL
| 2-8℃
|
| R777876B | 内源性过氧化物酶阻断剂（3% H₂O₂） | 6mL
| 2-8℃
|
| R777876C | Anti- Rabbit IgG（ HRP-Polymer） | 6mL
| 2-8℃
|
| R777876D | DAB显色试剂A（DAB×20倍浓缩液） | 2mL
| 2-8℃
|
| R777876E | DAB显色试剂B（H₂O₂×20倍浓缩液） | 2mL
| 2-8℃
|
| R777876F | DAB显色试剂C（TBS×20倍浓缩缓冲液） | 2mL
| 2-8℃
|
自备试剂及耗材：
1. 防脱玻片
2. PBS（pH7.2-7.6）
3. EDTA抗原修复液或枸橼酸盐缓冲液
使用说明：
A.石蜡切片热修复抗原染色程序:
1、制备切片，常规脱蜡至水。
2、3%H₂O₂，室温10 分钟以灭活内源性酶。蒸馏水洗2 分钟×3 次。
3、根据抗原抗体情况，必要时对切片进行抗原热修复或消化处理。
4、滴加封闭液,室温 10分钟。甩去多余液体, 不洗。
5、滴加适当稀释的一抗（兔 IgG），20-37 ℃ 1~2 小时或 4℃过夜。0.01M PBS 洗 2分钟×3次。（一抗的稀释度、孵育时间、温度与染色强度、背景有直接关系。一般来说，阳性染色强度不够时，可提高一抗浓度和延长孵育时间；背景过高时，可降低一抗浓度和缩短孵育时间。）
6、滴加聚合 HRP 标记抗兔IgG， 37℃孵育 30 分钟，PBS 或 TBS 冲洗，2 分钟×3 次。
7、DAB 显色:取1ml 蒸馏水,加试剂盒中A,B,C 试剂各1 滴,混匀后加至切片。室温显色, 镜下控制反应时间，一般5-30 分钟。蒸馏水洗涤。
8、 苏木素轻度复染、脱水,透明,封片、观察。
B.血涂片，细胞和冰冻切片染色程序
1.载玻片经防脱片剂处理（Poly-L-Lysine）。抗凝血经分层离心后涂片；培养细胞也可涂片或贴片生长；冰冻切片室温风扇吹干。
2.固定方案用4%多聚甲醛或丙酮固定10-20 分钟。
3.现配 3% H₂O₂ 与纯甲醇的混合液（体积比 1:4），室温浸泡样本 30 分钟，以灭活内源性过氧化物酶；结束后用蒸馏水洗涤 1–2 次。
其余步骤和石蜡切片3-8 步相同。
如果冰冻切片直接染色结果不理想时,可以参照石蜡切片对切片进行热修复，步骤和石蜡切片3-8 步相同。
本产品仅供科研使用。请勿用于医药、临床诊断或治疗，食品及化妆品等用途。
请勿存放于普通住宅区。 为了您的安全和健康，请穿好实验服并佩戴一次性手套和口罩操作。
(HRP-Polymer) Immunohistochemistry Kit (Rabbit) is a high-sensitivity immunohistochemistry kit. Horseradish peroxidase (HRP) is conjugated to the secondary antibody via polymer labeling to form large antibody-enzyme polymers, which replace the secondary and tertiary antibodies used in traditional methods. These polymers deliver potent signal amplification and excellent tissue and cell penetration, making them ideal for immunohistochemical applications. Compared with the conventional three-step method, this assay kit features simpler operation, faster testing, higher sensitivity and lower background staining.
Packing List:
| R777876 | Component | 1 Kit | Storage | | R777876A | Blocking Solution (5% BSA) | 6mL | 2-8℃ | | R777876B | Endogenous Peroxidase Blocker (3% H₂O₂) | 6mL | 2-8℃ | | R777876C | Anti-Rabbit IgG (HRP-Polymer) | 6mL | 2-8℃ | | R777876D | DAB Chromogenic Reagent A (20× Concentrate) | 2mL | 2-8℃ | | R777876E | DAB Chromogenic Reagent B (H₂O₂, 20× Concentrate) | 2mL | 2-8℃ | | R777876F | DAB Chromogenic Reagent C (TBS Buffer, 20× Concentrate) | 2mL | 2-8℃ |
|
Reagents and Consumables to Be Prepared Separately:
1. Adhesive glass slides
2. PBS solution (pH 7.2–7.6)
3. EDTA Antigen Retrieval Solution or Citrate Buffer
Instructions for Use:
A. Staining Protocol for Paraffin Sections with Heat-Induced Antigen Retrieval
1. Prepare sections and perform routine deparaffinization and rehydration.
2. Incubate sections in 3% H₂O₂ at room temperature for 10 minutes to inactivate endogenous enzymes. Rinse with distilled water 3 times, 2 minutes each time.
3. Perform heat-induced antigen retrieval or enzymatic digestion as required based on antigen-antibody characteristics.
4. Add blocking solution and incubate at room temperature for 10 minutes. Decant excess liquid without rinsing.
5. Add appropriately diluted primary antibody (Rabbit IgG). Incubate at 20–37 °C for 1–2 hours, or overnight at 4 °C. Rinse with 0.01 M PBS 3 times, 2 minutes each time.
(Note: The dilution ratio and incubation conditions of the primary antibody directly affect staining intensity and background. If positive staining is weak, increase antibody concentration or prolong incubation time. If high background occurs, reduce antibody concentration or shorten incubation time.)
6. Add HRP-polymer conjugated Anti-Rabbit IgG, and incubate at 37 °C for 30 minutes. Rinse with PBS or TBS 3 times, 2 minutes each time.
7. DAB Chromogenic Reaction: Add 1 drop each of Reagent A, B and C to 1 mL of distilled water, mix well and apply onto sections. Develop color at room temperature under microscopic observation for 5–30 minutes. Rinse thoroughly with distilled water.
8. Counterstain lightly with hematoxylin, followed by dehydration, clearing, mounting and microscopic examination.
B. Staining Protocol for Blood Smears, Cell Smears and Frozen Sections
1. Pre-treat glass slides with Poly-L-Lysine for anti-fall effect. Prepare smears from anticoagulated blood after density gradient centrifugation; prepare smears or adherent monolayers with cultured cells. Air-dry frozen sections at room temperature with a fan.
2. Fix samples with 4% paraformaldehyde or acetone for 10–20 minutes.
3. Prepare working solution by mixing 1 part 30% H₂O₂ with 50 parts pure methanol. Immerse slides in the mixture at room temperature for 30 minutes to block endogenous peroxidase activity. Rinse with distilled water 1–2 times.
Follow Steps 3 to 8 of the paraffin section protocol for the remaining operations.
If unsatisfactory results are obtained from direct staining of frozen sections, perform heat-induced antigen retrieval referring to the paraffin section procedure, and follow Steps 3 to 8 accordingly.
Disclaimer & Safety Instructions:
1. This product is for research use only. It shall not be used for medical treatment, clinical diagnosis, food, cosmetics or other related applications.
2. Do not store this product in residential areas. Wear lab coat, disposable gloves and face mask during operation for personal safety and health.
ZJ26F0636752
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