炭疽 (anthrax) 是由炭疽芽胞杆菌 (Bacillus anthracis) 引起的一种人兽共患急性传染病。主要发生于畜间，以牛、羊、马等草食动物最为易感。人主要通过接触患炭疽的动物或污染的动物制品、环境感染而患病。主要临床类型为皮肤炭疽，少数为肺炭疽和肠炭疽，可以继发败血症及脑膜炎。皮肤炭疽病死率较低，其他各型炭疽的病死率均较高。
炭疽杆菌是形体最大的革兰氏阳性杆菌，大小为1～1.5×3～5μm，两端平截，呈链状排列，镜下形态呈竹节状，链的长短因菌株和细菌所在环境而异。在人体、动物体内或特定环境条件下（于碳酸盐琼脂培养基上于5%～25% CO₂下培养），该菌可形成荚膜，印度墨汁染色可见杆菌周围的透明环，美蓝染色荚膜呈红色。在体外环境下可形成芽胞，未游离的芽胞在菌体中央。此菌无鞭毛，不运动。
细菌是否有芽孢以及芽孢的形状和位置都是细菌的重要特征，细菌的芽孢壁比营养细胞的细胞壁结构复杂、致密、通透性低，着色和脱色都比营养细胞困难。芽孢有很强的抗热和抗化学物质的特性，当采用碱性染料加热或延长染色时间时，使菌体和芽孢都能着色，再用蒸馏水冲洗脱去菌体颜色，但芽孢的颜色仍然可以保留，可以在显微镜下明显区分芽孢和营养体的形态。
阿拉丁荚膜染色液 (M'Fadyean法) 主要由美蓝染色液、次氯酸溶液组成，芽孢会被染成红色，菌体呈蓝色。本产品可以用于炭疽杆菌的病原学染色，操作简单，结果判断更可靠，但应注意，观察芽孢应在生物安全二级实验室中进行。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：
C1510420
| Component
| 2×50mL
| Storage
|
C1510420A
| 美蓝染色液
| 50mL
| RT
|
C1510420B
| 次氯酸溶液
| 50mL
| RT |
自备材料：
1、酒精灯、载玻片、接种环、显微镜
2、蒸馏水、实验样本 (皮肤溃疡的渗出物、排泄物、血、胸腹水及脑脊液等标本进行涂片或培养)
操作步骤(仅供参考)：
1、在洁净无油脂的载玻片上滴加1滴蒸馏水。
2、用接种环挑取少许实验样本，与蒸馏水混合。制成涂片，自然干燥。
3、用无水甲醇或乙醇固定涂片2~5min。
4、在涂液处滴加美蓝染色液，染色3~5min(如荚膜染色不清楚，可适当延长时间)。
5、用水轻轻冲洗玻片。
6、玻片入次氯酸溶液中脱色消毒2min，用水冲洗。
7、干燥后镜检，观察芽孢的形状、大小，在菌体中的位置以及是否使菌体胀大。
染色结果：
炭疽杆菌两端平齐、呈竹节状、长串联排列，荚膜为粉红色，菌体为蓝色。
注意事项：
1、玻片应洁净，无油污。
2、炭疽杆菌繁殖力、抵抗力与一般细菌相同，但芽胞抵抗力较强、危害性大，实验过程应在生物安全二级实验室中进行。
3、染色过程应小心操作，防止细菌脱落。
4、固定时不宜用高热火焰固定，常温固定即可。
5、实验冲洗后的废水以及吸水纸经高压灭菌处理或用次氯酸钠溶液处理过夜。
6、为了您的安全和健康，请穿实验服并戴一次性手套操作。
Anthrax is an acute zoonotic infectious disease caused by Bacillus anthracis. It mainly occurs in livestock, with herbivores such as cattle, sheep and horses being the most susceptible. Humans are mainly infected by contact with anthrax-affected animals, contaminated animal products or the polluted environment. The main clinical type is cutaneous anthrax, with a few cases of pulmonary anthrax and intestinal anthrax, which can be complicated by sepsis and meningitis. The mortality rate of cutaneous anthrax is relatively low, while that of other anthrax types is considerably higher.
Bacillus anthracis is the largest species of Gram-positive bacilli, with a size of 1–1.5×3–5 μm. It has truncated ends, arranges in chains, and presents a bamboo joint-like morphology under the microscope. The length of the chains varies depending on the bacterial strain and the surrounding environment. In the human or animal body, or under specific environmental conditions (cultured on carbonate agar medium with 5%–25% CO₂), the bacterium can form a capsule. A transparent ring around the bacilli can be observed by India ink staining, and the capsule appears red by methylene blue staining. The bacterium can produce spores in the external environment, with the non-free spores located in the center of the bacterial cells. This bacterium is non-flagellated and non-motile.
The presence of spores, as well as the shape and location of spores, are important characteristics of bacteria. The spore wall has a more complex, compact structure and lower permeability than the cell wall of vegetative cells, making it more difficult to stain and decolorize than vegetative cells. Spores have strong resistance to heat and chemical substances. When stained with basic dyes under heating or with extended staining time, both the bacterial cells and spores can be colored. Then, the color of the bacterial cells can be removed by rinsing with distilled water, while the color of the spores can be retained, allowing for clear morphological differentiation between spores and vegetative cells under a microscope.
Aladdin Capsule Stain Solution (M'Fadyean Method) is mainly composed of methylene blue staining solution and hypochlorous acid solution. With this solution, spores are stained red and bacterial cells are stained blue. This product can be used for the etiological staining of Bacillus anthracis, featuring simple operation and more reliable result interpretation. However, it should be noted that the observation of spores should be carried out in a biosafety level 2 (BSL-2) laboratory. This reagent is for research use only and is not suitable for clinical diagnosis or other purposes.
Product Components and Storage Conditions:
C1510420
| Component
| 2×50mL
| Storage
|
C1510420A
| Methylene Blue Staining Solution
| 50mL
| RT
|
C1510420B
| Hypochlorous Acid Solution
| 50mL
| RT |
Self-prepared Materials:
1. Alcohol lamp, glass slides, inoculation loops, microscopes
2. Distilled water, test samples (smears or cultures prepared from specimens such as exudates from skin ulcers, excreta, blood, pleural effusion, ascites and cerebrospinal fluid)
Operating Procedures (For Reference Only):
1. Place 1 drop of distilled water on a clean, grease-free glass slide.
2. Pick a small amount of the test sample with an inoculation loop, mix it thoroughly with the distilled water, prepare a smear, and let it air-dry naturally.
3. Fix the smear with anhydrous methanol or ethanol for 2–5 minutes.
4. Add methylene blue staining solution to cover the smear area, stain for 3–5 minutes (if the capsule staining is not clear, the staining time can be appropriately extended).
5. Rinse the glass slide gently with water.
6. Immerse the glass slide in hypochlorous acid solution for decolorization and disinfection for 2 minutes, then rinse it with water.
7. After drying, observe the slide under the microscope to check the shape, size, and position of spores in the bacterial cells, as well as whether the spores cause swelling of the bacterial cells.
Staining Results:
Bacillus anthracis has truncated ends, appears in a bamboo joint-like morphology and long chains arrangement; the capsule stains pink and the bacterial cells stain blue.
Precautions:
1. The glass slides should be clean and free of oil stains.
2. The reproductive capacity and resistance of Bacillus anthracis are similar to those of ordinary bacteria, but its spores have strong resistance and high hazard. The experiment should be carried out in a biosafety level 2 (BSL-2) laboratory.
3. Operate carefully during the staining process to prevent the bacteria from detaching from the slide.
4. Avoid high-temperature flame fixation during the fixation step; room-temperature fixation is sufficient.
5. The wastewater after rinsing in the experiment and the absorbent paper should be autoclaved or treated with sodium hypochlorite solution overnight.
6. For your safety and health, please wear a lab coat and disposable gloves during the operation.
ZJ26F0433798
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