Flow Cell Check
MinION: Protocol
Flow Cell Check V PQE_1004_v1_revAJ_06Jan2016
For Research Use Only
FOR RESEARCH USE ONLY
Contents
Overview
Running the flow cell check
概览
For Research Use Only
1. Introduction to the flow cell check
Purpose of the flow cell check
A Flow Cell Check is used to assess the number of nanopores that are available in the flow cell. It should be run when you are ready to use the flow cell, and within 3 months of purchase (except for Flongle flow cells, which currently have a shelf life of four weeks).
Oxford Nanopore Technologies will replace any flow cell that falls below the warranty number of active pores, provided the result is reported within two days of performing the Flow Cell Check and the storage recommendations have been followed.
| Flow cell | Minimum number of active pores covered by warranty |
|---|---|
| Flongle Flow Cell (FLO-FLG001) | 50 |
| Flongle Flow Cell (FLO-FLGOP1) | 30 |
| MinION/GridION Flow Cell | 800 |
| PromethION Flow Cell | 5000 |
Flow cell storage instructions
We recommend to store flow cells unopened, as follows:
| Flow cell | Storage instructions |
|---|---|
| Flongle Flow Cells | 2–8°C for four weeks |
| MinION/GridION Flow Cells | room temperature for one month, or at 2–8°C for 12 weeks |
| PromethION Flow Cells | 2–8°C for 12 weeks |
2. Flongle flow cell check
耗材
- Flongle device - flow cell and adapter
仪器
- MinION 或 GridION 测序仪
- Host computer connected to the Internet with MinKNOW and the Desktop Agent installed
重要
In this step, you will use the MinKNOW software to check the number of pores in your flow cell. This has to be done as soon as reasonably possible after receiving your Flongle flow cell. Flow cells can then be stored until required. If there is a delay between receiving and using the flow cells, another check should be performed before library preparation begins.
How it works
Flow cells are shipped with a QC DNA molecule present in the buffer. This molecule produces a distinctive nanopore signal. The MinKNOW™ software uses this signal to validate the integrity of the nanopore array before use and provides the user with an estimate of the number of simultaneously available channels for the experiment.
Place the Flongle adapter into the MinION or one of the five GridION positions.
The adapter should sit evenly and flat on the MinION Mk1B or GridION platform. This ensures the flow cell assembly is flat during the next stage.
重要
The adapter needs to be plugged into your device, and the device should be plugged in and powered on before inserting the Flongle flow cell.
Place the flow cell into the Flongle adapter, and press the flow cell down until you hear a click.
The flow cell should sit evenly and flat inside the adapter, to avoid any bubbles forming inside the fluidic compartments.
双击桌面上的MinKNOW图标，打开MinKNOW图形用户界面。
我们建议您使用纳米孔社区的账户信息登录MinKNOW软件。
您须连接到互联网方可登录。
用户可在此页面的右下角选择界面语言。
界面将提示您输入电邮地址和纳米孔密码：
如您在登录时遇到问题，请通过我们的纳米孔微信公众号（NanoporeSupport）或社区支持通道(https://community.nanoporetech.com/support) 联系我们的客服。与此同时，您可点击 Continue as guest ，以访客身份暂时使用MinKNOW。
Click the sequencing device connected to the computer.
The Sequencing Overview should show the flow cell has not had any checks carried out.
Below is an example of a GridION device with a Flongle inserted into position X1 and MinION Flow Cells in positions X2 - X5.
导航至“Start”（开始）页面，点击“Flow Cell Check”，打开测序芯片质检页面。
Choose the flow cell type from the drop down menu and fill in the flow cell ID.
Below is an example of a GridION device with a Flongle inserted into position X1.
重要
For Flongle, flow cell ID is not automatically assigned in MinKNOW. It is imperative the flow cell ID is entered in the correct format: ABC123 i.e. ([A-Z] x 3 [0-9] x 3).
The ID is case sensitive with no spaces. The MinKNOW software will not allow you to proceed until the flow cell ID has been entered correctly.
The user will be notified in the bottom right corner of the GUI if any information entered is in the wrong format or missing.
The Flongle flow cell ID is shown in the blue blox below:
Select 'Start' to begin the flow cell check.
随即进入“Sequencing Overview”（测序概览）页面。
测序芯片下方将有进度栏显示检测进度。
下图以Flongle配置测试芯片（CTC）为例（红框内为进度栏）。
The flow cell check should take a few minutes.
Flow cell health indicators
The quality of the flow cell will be shown as one of the three outcomes:
Yellow exclamation mark (Flongle flow cell) The number of sequencing pores is below warranty.
Green tick (MinION flow cell) The number of sequencing pores is above warranty.
Question mark (PromethION flow cell) A Flow Cell Check has not been run on the flow cell during this MinKNOW session.
Note: The indicator of quality (question mark, exclamation mark or tick) will only remain visible during a MinKNOW session. Once the MinKNOW service has ended, the status of the flow cell will be erased.
3. MinION Mk1C flow cell check
耗材
- MinION 和 GridION测序芯片
仪器
- MinION Mk1C
In this step, you will use the MinKNOW software to check the number of pores in your flow cell. This has to be done prior to loading your DNA or RNA library onto the flow cell.
How it works
Flow cells are shipped with a QC DNA molecule present in the buffer. This molecule produces a distinctive nanopore signal. The MinKNOW™ software uses this signal to validate the integrity of the nanopore array before use and provides the user with an estimate of the number of simultaneously available channels for the experiment.
Set up the MinION Mk1C and flow cell.
Power on the MinION Mk1C, as described in the MinION Mk1C protocol under Power.
Once successfully plugged in, you will see a light and hear the fan.
Open the MinION Mk1C lid and insert the flow cell under the clip.
Press down firmly to ensure good thermal and electrical contact.
Note: The above image uses a CTC for illustrative purposes. Please insert a flow cell.
我们建议您使用纳米孔社区的账户信息登录MinKNOW软件。
您须连接到互联网方可登录。
用户可在此页面的右下角选择界面语言。
界面将提示您输入电邮地址和纳米孔密码：
如您在登录时遇到问题，请通过我们的纳米孔微信公众号（NanoporeSupport）或社区支持通道(https://community.nanoporetech.com/support) 联系我们的客服。与此同时，您可点击 Continue as guest ，以访客身份暂时使用MinKNOW。
Select the Mk1C in the connection manager to open the device.
Navigate to the Start page and select 'Flow Cell Check' to open the flow cell check page.
When you see the flow cell type and flow cell IDs recognised, click 'Start' to begin.
The user will be automatically navigated to the Sequencing Overview page.
Run state can be viewed here and a progress bar is displayed at the base of the flow cell. 
Number of pores available will be displayed here once check is completed. 
The flow cell check should take a few minutes.
Flow cell health indicators
The quality of the flow cell will be shown as one of the three outcomes on the Sequencing Overview page:
Yellow exclamation mark: The number of sequencing pores is below warranty.
Green tick: The number of sequencing pores is above warranty and ready for sequencing.
Question mark: A flow cell check has not been run on the flow cell during this MinKNOW session.
Note: the indicator of quality will only remain visible during the MinKNOW session when testing occurred. Once the sequencing device has been re-started, the status of the flow cell will be erased.
4. MinION Mk1B/GridION flow cell check
耗材
- MinION 和 GridION测序芯片
仪器
- MinION Mk1B/GridION
- Host computer connected to the Internet with MinKNOW and the Desktop Agent installed
In this step, you will use the MinKNOW software to check the number of pores in your flow cell. This has to be done prior to loading your DNA or RNA library onto the flow cell.
How it works
Flow cells are shipped with a QC DNA molecule present in the buffer. This molecule produces a distinctive nanopore signal. The MinKNOW™ software uses this signal to validate the integrity of the nanopore array before use and provides the user with an estimate of the number of simultaneously available channels for the experiment.
Open the MinION Mk1B/GridION lid and insert the MinION Flow Cell.
Insert the flow cell in the MinION Mk1B/GridION by sliding the flow cell under the clip. Firmly press down on the flow cell to ensure good thermal and electrical contact.
Connect the assembled MinION Mk1B and flow cell to the host computer.
Figure SSFC: Assembled MinION Mk1B and MinION SpotON Flow Cell connected to through the USB 3.0 port to the host computer
Once successfully plugged in, you will see a light and hear the fan.
双击桌面上的MinKNOW图标，打开MinKNOW图形用户界面。
我们建议您使用纳米孔社区的账户信息登录MinKNOW软件。
您须连接到互联网方可登录。
用户可在此页面的右下角选择界面语言。
界面将提示您输入电邮地址和纳米孔密码：
如您在登录时遇到问题，请通过我们的纳米孔微信公众号（NanoporeSupport）或社区支持通道(https://community.nanoporetech.com/support) 联系我们的客服。与此同时，您可点击 Continue as guest ，以访客身份暂时使用MinKNOW。
Click the sequencing device connected to the computer.
The Sequencing Overview should show the flow cell has not had any checks carried out.
Below is an example of a GridION device with a Flongle inserted into position X1 and MinION Flow Cells in positions X2 - X5.
导航至“Start”（开始）页面，点击“Flow Cell Check”，打开测序芯片质检页面。
When you see the flow cell type and flow cell IDs recognised, click 'Start' to begin.
随即进入“Sequencing Overview”（测序概览）页面。
测序芯片下方将有进度栏显示检测进度。
下图以Flongle配置测试芯片（CTC）为例（红框内为进度栏）。
The flow cell check should take a few minutes.
测序芯片健康状况指示框
测序概览（Sequencing Overview）页面会显示测序芯片的质量。质量分为三个等级：
黄色!（图中Flongle测序芯片）: 活性测序孔数量低于质保范围。将芯片从测序设备中取出后重新插入，并运行测序芯片质检。如果测序芯片活性孔数仍然低于质保范围，请您发邮件至support@nanoporetech.com。
绿色√（图中MinION 测序芯片）： 活性测序孔数量在质保范围之上。
?（图中PromethION 测序芯片）： 测序芯片在本次MinKNOW使用期间还未经过质检。
请注意： 芯片质量指示框仅在当次MinKNOW使用期间可见。一旦该次使用结束，测序芯片的状态将被清除。
5. PromethION flow cell check
耗材
- PromethION 测序芯片
仪器
- PromethION测序设备
How it works
Flow cells are shipped with a QC DNA molecule present in the buffer. This molecule produces a distinctive nanopore signal. The PromethION software uses this signal to validate the integrity of the nanopore array before use and provides the user with an estimate of the number of simultaneously available channels for the experiment. Active pores are reported in four groups, each of which may be used in turn when running long experiments e.g. a 64 hour sequencing run. Shorter experiments may not use all groups.
重要
将芯片从冰箱中取出后，请将其置于室温环境孵育20分钟再插入PromethION测序仪。潮湿环境下的测序芯片上可能会形成冷凝水。因此，请检查测序芯片顶部和底部的金色连接器引脚处是否有水凝结。如有，请使用无纤维布擦干。请确保测序芯片底部有热垫（黑色）覆盖。
将待运行的测序芯片插入PromethION上相应插槽的对接端口。
将芯片连接器应完全插入对接端口。请确保用于固定芯片的卡夹卡入原位。
重要
正确插入PormethION测序芯片
将测序芯片与连接器横竖对齐后，即可轻松插入至测序设备的相应位置。
用力下压芯片至插槽，并确认卡夹位置归位。
如插入配置测试芯片的角度出现偏差，可能会损坏PromethION上的引脚并影响测序结果。如您发现PromethION测序仪芯片卡槽上的引脚损坏，请通过电子邮件（Support@nanoporetech.com）联系我们的客服。 
测序芯片正确插入 PromethION 后，将自动运行硬件检测：
- 绿色√：测序芯片通过硬件检测
- 红色√：请重新插入测序芯片
双击桌面上的MinKNOW图标，打开MinKNOW图形用户界面。
我们建议您使用纳米孔社区的账户信息登录MinKNOW软件。
您须连接到互联网方可登录。
用户可在此页面的右下角选择界面语言。
界面将提示您输入电邮地址和纳米孔密码：
如您在登录时遇到问题，请通过我们的纳米孔微信公众号（NanoporeSupport）或社区支持通道(https://community.nanoporetech.com/support) 联系我们的客服。与此同时，您可点击 Continue as guest ，以访客身份暂时使用MinKNOW。
Click the sequencing device connected to the computer.
The Sequencing Overview page should show the flow cell has not had any checks carried out.
导航至“Start”（开始）页面，点击“Flow Cell Check”，打开测序芯片质检页面。
当界面显示测序芯片类型并识别出芯片ID时，点击“Start”（开始）启动芯片质检。
随即进入“Sequencing Overview”（测序概览）页面。
测序芯片下方将有进度栏显示检测进度。
下图以Flongle配置测试芯片（CTC）为例（红框内为进度栏）。
The flow cell check should take a few minutes.
测序芯片健康状况指示框
测序概览（Sequencing Overview）页面会显示测序芯片的质量。质量分为三个等级：
黄色!（图中Flongle测序芯片）: 活性测序孔数量低于质保范围。将芯片从测序设备中取出后重新插入，并运行测序芯片质检。如果测序芯片活性孔数仍然低于质保范围，请您发邮件至support@nanoporetech.com。
绿色√（图中MinION 测序芯片）： 活性测序孔数量在质保范围之上。
?（图中PromethION 测序芯片）： 测序芯片在本次MinKNOW使用期间还未经过质检。
请注意： 芯片质量指示框仅在当次MinKNOW使用期间可见。一旦该次使用结束，测序芯片的状态将被清除。