病毒性肝炎是由肝炎病毒引起的肝实质细胞变性坏死为主要病变的传染病，乙型肝炎病毒是世界范围内传播的乙肝病毒，HBsAg存在于肝细胞质内。目前可通过免疫组化方法显示乙型肝炎病毒的HBsAg和HBcAg，显示HBsAg的方法有Shikata地衣红法、醛品红法、维多利亚蓝法等。
阿拉丁乙型肝炎病毒染色液 (Shikata地衣红法) 操作简单、特异性高，但染色效果可能受到不同批次原料而有差异，其可能机制是地衣红与HBsAg内胱氨酸的双硫键结合，进而显色。该试剂仅用于科研领域，不适用于临床诊断或其他用途。
产品组分及储存条件：
H1510444
| Component
| 3×50mL
| Storage
|
H1510444A
| Shikata 氧化剂
| 25mL
| RT. Store in the dark.
|
H1510444B
| Shikata 酸溶液
| 25mL
| RT
|
H1510444C
| Shikata 漂白液
| 50mL
| RT
|
H1510444D
| 酸性地衣红染色液
| 50mL
| 4°C. Store in the dark. |
临用前，取A、B等量混合，即为Shikata 酸化液，即配即用。
自备材料：
10%福尔马林固定液、蒸馏水、系列乙醇
操作步骤(仅供参考)：
1、组织固定于10%的中性福尔马林固定液，常规脱水包埋。
2、切片厚4μm，常规二甲苯或脱蜡透明液脱蜡至水。
3、入Shikata酸化液氧化5min，稍水洗。
4、入Shikata漂白液漂白2min，脱去酸化液的着色，流水冲洗2min。
5、入70%乙醇稍浸洗。入酸性地衣红染色液加盖浸染5h或更长时间。
6、二甲苯或脱蜡透明液透明，中性树胶封固。
染色结果：
HBsAg 阳性物质
| 棕红色至深棕色
|
背景
| 不同程度的淡红色
|
注意事项：
1、Shikata酸化液必须即配即用，混合后久置会降低其氧化力。
2、酸性地衣红染色液临用前应恢复至室温，染液可重复使用数次。
3、染色5h后，可取出切片在70%乙醇中浸洗一下，再入蒸馏水水洗，镜下观察；如染色较淡，可在70%乙醇中浸洗后入酸性地衣红染色液染色，必要时可过夜染。
4、地衣红染色时，脂褐素亦可呈阳性反应；其区别方法是：脂褐素着色较深，在细胞质内呈大小均匀的小颗粒；HBsAg着色较浅，呈细微颗粒状。
5、为了您的安全和健康，请穿实验服并戴一次性手套操作。
6、试剂开封后请尽快使用，以防影响后续实验效果。
Viral hepatitis is an infectious disease characterized by degeneration and necrosis of liver parenchymal cells, which is caused by hepatitis viruses. Hepatitis B virus (HBV) is a globally prevalent hepatitis virus, and hepatitis B surface antigen (HBsAg) is present in the cytoplasm of hepatocytes. At present, immunohistochemical methods can be used to detect HBsAg and hepatitis B core antigen (HBcAg) of HBV. The methods for detecting HBsAg include Shikata’s orcein method, aldehyde-fuchsin method, Victoria blue method, etc.
Aladdin Hepatitis B Virus Staining Solution (Shikata’s Orcein Method) features simple operation and high specificity, but the staining effect may vary with different batches of raw materials. The potential mechanism is that orcein binds to the disulfide bonds of cystine in HBsAg, thereby producing color development. This reagent is intended for research use only and is not suitable for clinical diagnosis or any other purposes.
Product Components and Storage Conditions:
H1510444
| Component
| 3×50mL
| Storage
|
H1510444A
| Shikata Oxidant
| 25mL
| RT. Store in the dark.
|
H1510444B
| Shikata Acid Solution
| 25mL
| RT
|
H1510444C
| Shikata Bleaching Solution
| 50mL
| RT
|
H1510444D
| Acidic Orcein Staining Solution
| 50mL
| 4°C. Store in the dark. |
Prior to use, mix equal volumes of Solution A and Solution B to prepare the Shikata Acidified Solution. The solution should be prepared fresh and used immediately.
Materials Required (Self-prepared):
10% formalin fixative, distilled water, graded ethanol solutions
Operating Procedures (For Reference Only):
1. Fix the tissue in 10% neutral formalin fixative, followed by routine dehydration and embedding.
2. Cut into sections with a thickness of 4 μm, then perform routine dewaxing to water using xylene or dewaxing clearing solution.
3. Immerse the sections in Shikata acidified solution for oxidation for 5 minutes, then rinse briefly with water.
4. Transfer the sections into Shikata bleaching solution for bleaching for 2 minutes to remove the stain from the acidified solution, followed by rinsing under running water for 2 minutes.
5. Rinse the sections slightly in 70% ethanol. Then place them in acidic orcein staining solution, cover the container, and stain for 5 hours or longer.
6. Clear the sections with xylene or dewaxing clearing solution, and mount the sections with neutral balsam.
Staining Results:
HBsAg-positive Substances
| Reddish-brown to dark brown
|
Background
| Pale red of varying degrees
|
Precautions:
1. Shikata acidified solution must be prepared fresh and used immediately. Prolonged storage after mixing will reduce its oxidizing capacity.
2. Bring the acidic orcein staining solution to room temperature before use. The staining solution can be reused several times.
3. After 5 hours of staining, the sections can be removed, rinsed briefly in 70% ethanol, and then washed with distilled water for microscopic examination. If the staining is faint, rinse the sections in 70% ethanol and then restain them in acidic orcein staining solution; overnight staining is permissible if necessary.
4. Lipofuscin may also yield a positive reaction with orcein staining. The distinguishing features are as follows: lipofuscin stains darker and appears as uniformly sized fine granules in the cytoplasm; whereas HBsAg stains lighter and presents as fine granular deposits.
5. For your safety and health, wear a lab coat and disposable gloves during the operation.
6. Please use the reagents as soon as possible after opening to avoid compromising subsequent experimental results.
ZJ26F0333229
危险品化学品经营许可证（带存储）