人成纤维细胞-成人心室

【人成纤维细胞-成人心室】下面是细胞株培养的具体无菌技术要点：

1. 实验开始前，无菌室和无菌操作台需要紫外光照射30到60分钟完全灭菌，用70%ethanol擦拭无菌操作台面，并且打开无菌操作台风扇运行10分钟之后，才可以进行实验操作。每次操作只能处理一株细胞株，即使培养基完全相同也不可共用培养基，以避免细胞间污染或失误混淆。实验完成后，请将实验物品拿出工作台，用70%ethanol再次擦拭无菌操作台面。操作间隔应该让无菌操作台运转10分钟到15分钟后，才能进行下一个细胞株的操作。

2. 无菌操作工作的区域应保持清洁和宽敞，必要物品（试管架、吸管、吸取器或吸管盒等）可以暂时放置，其它实验用品使用完毕后应移出，有利于空气流通。实验用品需70%ethanol擦拭后方可带入无菌操作台内。实验操作过程应在台面的中央无菌区域内完成，请勿在边缘非无菌区域进行操作。

3. 小心取用无菌实验物品，避免细菌污染。请勿触碰吸管尖头部和容器瓶口处，也不要在打开的容器正上方进行操作。容器打开后，请用手夹住瓶盖并轻握瓶身，倾斜大约45°角取用，尽量不要将瓶盖口朝上放置于桌面。

4. 工作人员应当首先注意自身安全，必须穿戴齐实验衣和实验手套后才能进行实验。对于病毒感染或是来自人类的细胞株应当特别小心操作，并选择适当等级的无菌操作台进行（至少是Class II）。操作过程中，应避免引起aerosol的产生，小心有毒性药品，例如DMSO和TPA等，并避免针头的伤害等等。

5. 定期检测下列项目：

5.1. CO2钢瓶的CO2压力

5.2. CO2培养箱的CO2浓度、温度、和水盘是否有污染（水盘的水需用无菌水，每周定时更换）。

5.3. 无菌操作台内的airflow压力，定期更换紫外线灯管和HEPA过滤膜，预滤网（300小时/预滤网，3000小时/HEPA）。

6. 水槽内可添加消毒剂（Zephrin 1:750），需定期更换水槽中的水。

【人成纤维细胞-成人心室】齐一生物友情提醒，本产品仅供科研使用，不得用于临床及诊断使用！

一细胞说明，具体详细请见细胞附带说明书

 二、客户自备试剂

1、PBS  

2、 2、RPMI-1640 培养基（不含 Hepes）+ 10%胎牛血清+ 1%双抗

3、0.25%  （W/V）胰蛋白酶（含 0.02% EDTA）

三、细胞背景

1、生长方式：贴壁

2、种属：Homo sapiens  人

四、培养条件

1、培养基：RPMI-1640 培养基（不含 Hepes）+ 10%胎牛血清+ 1%双抗

2、温度：37.0°C

3、气体:空气  95%，CO2 5%

【人成纤维细胞-成人心室】五、培养方法

 收到细胞后，在倒置显微镜下观察整个细胞生长情况：

 如果细胞未长满，用 75%酒精喷洒整个瓶消毒后放到超菌台内，严格无菌操作，打开细胞培养瓶，留10ml 培养液继续培养。如果细胞已长满（达 80-90%）。即可进行传代，具六、体步骤如下：

1，弃去培养液，用PBS洗 1-2次。

2，向瓶内加入 1.0-2.0ml 胰蛋白酶液，在倒置显微镜下观察细胞消化情况，若细胞大部分变圆，迅速拿回操作台，吸取胰蛋白酶，加含有 6ml  含 10%血清的培养液，轻轻吹打细胞。

3，加入等量的的培养液，轻轻吹打混匀后吸出一半，分到新的培养

4，传代比例：1:2-1:3  

七、常见问题及解决方案

1、培养瓶有破裂，培养液有漏液：细胞极大可能会污染，所以我们会及时安排帮老师解决。

2、细胞漂浮：培养瓶不开封，瓶口酒精擦拭后平躺放置在培养箱。次日观察，如细胞大部分又贴回瓶底，表明细胞活力正常，剩余漂浮的细胞可以去掉，留 10ml 培养液培养观察，细胞生长至汇合度 80%，进行消化传代；如细胞还是不贴壁，将细胞离心收集转到新培养瓶，原培养瓶加部分培养液继续培养，中间注意观察，我们的技术人员会一直跟踪指导，直到问题解决。

细胞培养代理品牌

细胞培养代理品牌

【人成纤维细胞-成人心室】齐一生物科技（上海）有限公司是一家专业生产和销售各种生化试剂,细胞株、原代细胞、菌种、医药中间体,标准品和对照品的大型化学科技公司. 齐一生物销售：021－6034 8496；181214 53965；173021 04490；Web:www.qiyibio.com

9241768QIAxtractor, Basic AgreementThe Basic Cover Agreement, QIAxtractor provides full service coverage for the QIAxtractor ａnd the CAS 1280. Costs for service visits are fully covered including travel, labor, ａnd parts. One on-site pipetting head service ａnd inspection is included. With a Basic Cover Agreement, there is a response time of 5 working days.CNY

9241656Transfer Rack Plate, APCNY

9241630Shaker Adjustment Kit, APCNY

9241367Damper, door, EZ1aCNY

9240759Update, Sliding Door Latch, AP98CNY

9243548Pyro Q96 ID, PM Service"Service response time (repair): not applicable

Periodic inspection/maintenance: yes

Inspection/maintenance delivery: onsite

Cost coverage for Repair Parts : not included

Cost coverage for labor: yes, full coverage

Cost coverage for travel: yes, full coverage

Replacement system (Loaner) provision: not applicable

Transportation cost coverage: yes, full coverage"CNY

9242766Cogwheel, x-drive, portal, QsymCNY

9242710Validation Support, 2d, QIAgilityThe QIAGEN QIAgility Guided Validation Support service includes all travel ａnd labor charges for a site visit up to 2 days by a QIAGEN Specialist.  A QIAGEN specialist will provide an introduction to the QIAgility system ａnd software, review the User Manual, ａnd practical hands-on operation. The Guided Validation service includes experimental design consultation, internal validation documentation support, ａnd QIAGEN sponsored data templates.  Enough consumable chemistry ａnd accessories (QIAGEN or other vendors) should be purchased to cover the validation experimental design ａnd are not included in the cost of validation support. Please indicate which processes (quant setup, normalization, PCR setup, CE setup) ａnd kit manufacturer (Applied Biosystems, Promega, QIAGEN) need to be included in the validation.CNY

9242253Cable, x, portal AS, QsymCNY

9242234RGQ, IQ/OQ Service"Instrument qualification support service for the Rotorgene Q ａnd Rotorgene 6000. The product contains the IQ/OQ documentation ａnd the performance of the qualification protocols (travel ａnd labor costs included). 

During the service, the compliance with the manufacturer＇s installation requirements is testified ａnd the performance verification & dynamic calibration of the Rotorgene Q is performed."CNY

9242172Repair fee, Depot Repair, TissueLyser IICNY

5000-1403Bumpers Side, VTX2CNY

5000-1401Guide Rack Back Asmbly, VTX2CNY

5000-1311Capacitor 8UF, SKR1CNY

5000-1273Door Damper Assembly, DML2CNY

5000-1258Y Optical Sensor CMPLT, DML2CNY

5050-1074 V.1CD Rom Manual - LumiCheck PlateCNY

5050-1069 V.1CD Rom Manual - Rotary Shaker I, 240VCNY

6000-5001Costar Plate LidsCNY

6000-3120Plate Washer I - WasteReserv.CapAssemblyCNY

5000-1420Driveplate Asmly 099A, VTX2CNY

5127-1220hc2 Sample Conversion KitCNY

9243701careHPV, TrainingCNY

EA11423EASYartus BKV RG_ICNY

800190Sequencing in a Box ServiceCNY

800110Single Read, w/ prep"Bacterial clone cultivation ａnd template purification or PCR product purification, including DNA quality control

Repetition of sequencing, if necessary

Data collected to maximal read length"CNY

132538NeXtal Stock Dioxane (50)100% v/v Dioxane - 50mL (5 x 10 ml tube)CNY

338162RT2 PCR Array Loading ReservoirCNY

338114Mouse XpressRef Universal Total RNACNY

203743QIAGEN Fast Cycling PCR Kit (200)For 200 x 20 µl reactions: 2 x 1 ml Fast Cycling Master Mix, 2x (containing dNTPs, patent pending buffer composition), RNase-free water, 10 x CoralLoad Fast Cycling Dye, 5 x Q-SolutionCNY