Add Total Protein Normalization to ECL or NIR Western Blots
Azure Biosystems provides a custom workflow for Total Protein Staining (TPS). The wider linearity and dynamic range of TPS is the new standard preferred by journals for accurate Western Blot quantification. We offer three imaging systems with optional fluorescent channels.
The Azure Q Module is an optional fluorescence channel intended for detecting TPS, such as TotalStain Q. With an extra channel, you are now able to detect up to two proteins of interest + total protein.
Normalize signal in your chemiluminescent western blots with the addition of a fluorescent channel. Detect more than just housekeeping proteins for quantitative western blots on top of your traditional ECL detection.
Near-infrared lasers provide the highest signal to noise ratio
with minimal crosstalk. The addition of a third fluorescent
channel to the camera-based imager allows the simultaneous
detection of housekeeping proteins or a total protein stain
The new Sapphire FL supports a broad range of excitation and emission wavelengths. We recommend the standard 532 optical module for detection of TotalStain Q.
TotalStain Q is a total protein staining reagent, compatible with chemiluminescent or near infrared fluorescent Western Blot detection. The linear range between 1–50 μg of lysate makes it ideal for normalization of low expressed proteins that require high loads of protein to reach the sensitivity for the detection of the target.
Quantitative Western blots require the normalization of signal to an internal loading control. Guidelines from leading journals state that total protein normalization is preferred over normalization to housekeeping proteins.
Journal of Biological Chemistry
Transparency is the Key to Quality
“It is not uncommon to “correct” Western blot signals for protein loading by normalizing to [housekeeping proteins present in the same replicate], e.g.: β-actin, α-tubulin, transferrin, GAPDH, HPRT1. The problem with this approach is that a linear relationship between signal intensity and the mass or volume of sample loaded must be confirmed for every antigen… It is typically better to normalize Western blots using total protein loading as the denominator [given its wider linear range].”.
Fosang, Amanda J, and Roger J Colbran. “Transparency Is the Key to Quality.” The Journal of biological chemistry vol. 290,50 (2015): 29692-4. doi:10.1074/jbc.E115.000002
Journal of Proteomics
Tubulin or Not Tubulin: Heading Toward Total Protein Staining as Loading Control in Western Blots
“Total protein staining (TPS) represents the actual loading amount more accurately than HKPs due to minor technical and biological variation. Further, the broad dynamic range of TPS solves the issue of HKPs that commonly fail to show loading differences above small loading amounts of 0.5-10 μg.”
Moritz CP. Tubulin or Not Tubulin: Heading Toward Total Protein Staining as Loading Control in Western Blots. Proteomics. 2017 Oct;17(20). doi: 10.1002/pmic.201600189. PMID: 28941183
Document Type | Description | |
---|---|---|
Application Note | Three-Color Western Blots with AzureSpectra Antibodies | DOWNLOAD |
Application Note | Imaging Three-color Western Blots with the Azure c600 | DOWNLOAD |
Application Note | Accurate Western Blot Normalization with AzureRed Fluorescent Protein Stain | DOWNLOAD |
Application Note | Phosphorylated Protein Detection is More Efficient by Fluorescent Western Blot | DOWNLOAD |
Application Note | Increasing Assay Efficiency with Four Color Detection | DOWNLOAD |
Brochure | The Azure Q Module | DOWNLOAD |
Brochure | Sapphire Applications Overview | DOWNLOAD |
Brochure | Sapphire FL Biomolecular Imager | DOWNLOAD |
Cookie | Duration | Description |
---|---|---|
cookielawinfo-checbox-analytics | 11 months | This cookie is set by GDPR Cookie Consent plugin. The cookie is used to store the user consent for the cookies in the category "Analytics". |
cookielawinfo-checbox-functional | 11 months | The cookie is set by GDPR cookie consent to record the user consent for the cookies in the category "Functional". |
cookielawinfo-checbox-others | 11 months | This cookie is set by GDPR Cookie Consent plugin. The cookie is used to store the user consent for the cookies in the category "Other. |
cookielawinfo-checkbox-advertisement | 1 year | Set by the GDPR Cookie Consent plugin, this cookie is used to record the user consent for the cookies in the "Advertisement" category . |
cookielawinfo-checkbox-necessary | 11 months | This cookie is set by GDPR Cookie Consent plugin. The cookies is used to store the user consent for the cookies in the category "Necessary". |
cookielawinfo-checkbox-performance | 11 months | This cookie is set by GDPR Cookie Consent plugin. The cookie is used to store the user consent for the cookies in the category "Performance". |
CookieLawInfoConsent | 1 year | Records the default button state of the corresponding category & the status of CCPA. It works only in coordination with the primary cookie. |
elementor | never | This cookie is used by the website's WordPress theme. It allows the website owner to implement or change the website's content in real-time. |
JSESSIONID | session | The JSESSIONID cookie is used by New Relic to store a session identifier so that New Relic can monitor session counts for an application. |
LS_CSRF_TOKEN | session | Cloudflare sets this cookie to track users’ activities across multiple websites. It expires once the browser is closed. |
viewed_cookie_policy | 11 months | The cookie is set by the GDPR Cookie Consent plugin and is used to store whether or not user has consented to the use of cookies. It does not store any personal data. |
woolentor_already_views_count_product | session | Woolentor set this cookie to store product count in cart. |
woolentor_viewed_products_list | 5 days | Woolentor set this cookie to store products in a cart. |
zfccn | session | Zoho sets this cookie for website security when a request is sent to campaigns. |
Cookie | Duration | Description |
---|---|---|
_zcsr_tmp | session | Zoho sets this cookie for the login function on the website. |
S | 1 hour | Used by Yahoo to provide ads, content or analytics. |
Cookie | Duration | Description |
---|---|---|
_uetsid | 1 day | Bing Ads sets this cookie to engage with a user that has previously visited the website. |
_uetvid | 1 year 24 days | Bing Ads sets this cookie to engage with a user that has previously visited the website. |
gaconnector_page_visits | 6 months | This cookie is set to collect information on user behavior and navigation which is used to optimize the website. |
gaconnector_pages_visited_list | 6 months | This cookie is set to collect information on user behavior and navigation which is used to optimize the website. |
SRM_B | 1 year 24 days | Used by Microsoft Advertising as a unique ID for visitors. |
Cookie | Duration | Description |
---|---|---|
_ga | 2 years | The _ga cookie, installed by Google Analytics, calculates visitor, session and campaign data and also keeps track of site usage for the site's analytics report. The cookie stores information anonymously and assigns a randomly generated number to recognize unique visitors. |
_ga_* | 1 year 1 month 4 days | Google Analytics sets this cookie to store and count page views. |
_ga_B687YMESCW | 2 years | This cookie is installed by Google Analytics. |
_ga_devsite | 2 years | This cookie is installed by Google Analytics. |
_gat_gtag_UA_* | 1 minute | Google Analytics sets this cookie to store a unique user ID. |
_gat_gtag_UA_43984149_1 | 1 minute | Set by Google to distinguish users. |
_gcl_au | 3 months | Provided by Google Tag Manager to experiment advertisement efficiency of websites using their services. |
_gid | 1 day | Installed by Google Analytics, _gid cookie stores information on how visitors use a website, while also creating an analytics report of the website's performance. Some of the data that are collected include the number of visitors, their source, and the pages they visit anonymously. |
0d269e8747 | session | Zoho PageSense sets this cookie to evaluate visitor, session and campaign data and keeps track of site usage for the site’s analytics report and is set by Zoho Analytics. |
cb_anonymous_id | 1 year | Clearbit sets this cookie to track page views and traits for Clearbit. |
cb_group_id | 1 year | Clearbit sets this cookie to track page views and traits for Clearbit. |
CONSENT | 2 years | YouTube sets this cookie via embedded youtube-videos and registers anonymous statistical data. |
gaconnector_all_traffic_sources | 6 months | This cookie determines how the user accessed the website. This information is used by the website operator to measure the efficiency of their marketing. |
gaconnector_browser | 6 months | This cookie is set to determine the browser. |
gaconnector_city | 6 months | This cookie is set to determine the city. |
gaconnector_country | 6 months | This cookie is set to determine the country. |
gaconnector_country_code | 6 months | This cookie is set to determine the country code. |
gaconnector_device | 6 months | This cookie is set to determine the device. |
gaconnector_fc_campaign | 6 months | This cookie is set to determine the first click campaign. |
gaconnector_fc_channel | 6 months | This cookie is set to determine the first click channel. |
gaconnector_fc_content | 6 months | This cookie is set to determine the first click content. |
gaconnector_fc_landing | 6 months | This cookie is set to determine the first click landing page. |
gaconnector_fc_medium | 6 months | This cookie is set to determine the first click medium. |
gaconnector_fc_referrer | 6 months | This cookie is set to determine the first click referrer. |
gaconnector_fc_source | 6 months | This cookie is set to determine the first click source. |
gaconnector_fc_term | 6 months | This cookie is set to determine the first click term. |
gaconnector_GA_Client_ID | 6 months | This cookie stores the Google Analytics Client ID. |
gaconnector_gclid | 6 months | This cookie is set by GA Connector service to store the Google Click Identifier. |
gaconnector_ip_address | 6 months | This cookie is set to determine the IP address. |
gaconnector_latitude | 6 months | This cookie is set to determine the latitude. |
gaconnector_lc_campaign | 6 months | This cookie is set to determine the last click campaign. |
gaconnector_lc_channel | 6 months | This cookie is set to determine the last click channel. |
gaconnector_lc_content | 6 months | This cookie is set to determine the last click content. |
gaconnector_lc_landing | 6 months | This cookie is set to determine the last click landing page. |
gaconnector_lc_medium | 6 months | This cookie is set to determine the last click medium. |
gaconnector_lc_referrer | 6 months | This cookie is set to determine the last click referrer. |
gaconnector_lc_source | 6 months | This cookie is set to determine the last click source. |
gaconnector_lc_term | 6 months | This cookie is set to determine the last click term. |
gaconnector_longitude | 6 months | This cookie is set to determine the longitude. |
gaconnector_OS | 6 months | This cookie is set to determine the operating system. |
gaconnector_region | 6 months | This cookie is set to determine the region. |
gaconnector_time_passed | 6 months | This cookie stores data on the time spent on the website and its sub-pages, during the current session. |
gaconnector_time_zone | 6 months | This cookie is set to determine the time zone. |
gaconnector2_id | never | This cookie is used by the GA Connector service to gather information regarding visitor activity and related advertising campaigns on the website upon enquiry. |
MR | 7 days | This cookie, set by Bing, is used to collect user information for analytics purposes. |
zabUserId | 1 year | Zoho PageSense sets this cookie to store a unique ID for every user who visits that website. |
Cookie | Duration | Description |
---|---|---|
ANONCHK | 10 minutes | The ANONCHK cookie, set by Bing, is used to store a user's session ID and also verify the clicks from ads on the Bing search engine. The cookie helps in reporting and personalization as well. |
cb_user_id | 1 year | Clearbit sets this cookie to collect data on visitors. This information is used to assign visitors into segments, making website advertising more relevant. |
IDE | 1 year 24 days | Google DoubleClick IDE cookies are used to store information about how the user uses the website to present them with relevant ads and according to the user profile. |
MUID | 1 year 24 days | Bing sets this cookie to recognize unique web browsers visiting Microsoft sites. This cookie is used for advertising, site analytics, and other operations. |
NID | 6 months | NID cookie, set by Google, is used for advertising purposes; to limit the number of times the user sees an ad, to mute unwanted ads, and to measure the effectiveness of ads. |
test_cookie | 15 minutes | The test_cookie is set by doubleclick.net and is used to determine if the user's browser supports cookies. |
VISITOR_INFO1_LIVE | 5 months 27 days | A cookie set by YouTube to measure bandwidth that determines whether the user gets the new or old player interface. |
YSC | session | YSC cookie is set by Youtube and is used to track the views of embedded videos on Youtube pages. |
yt-remote-connected-devices | never | YouTube sets this cookie to store the video preferences of the user using embedded YouTube video. |
yt-remote-device-id | never | YouTube sets this cookie to store the video preferences of the user using embedded YouTube video. |
yt.innertube::nextId | never | This cookie, set by YouTube, registers a unique ID to store data on what videos from YouTube the user has seen. |
yt.innertube::requests | never | This cookie, set by YouTube, registers a unique ID to store data on what videos from YouTube the user has seen. |
Cookie | Duration | Description |
---|---|---|
__rafm | session | No description available. |
__rasesh | 2 years 9 months 23 days | No description available. |
__tld__ | session | Description is currently not available. |
_bioz_depot_session | session | Description is currently not available. |
_depot_session | session | No description |
_vh_gid | 7976 years 10 months 22 days 20 hours 2 minutes | No description |
_vh_sid | 15 minutes | No description |
_vh_sid_pn | session | No description |
383aeadb58 | session | No description available. |
53d772c94a | session | Description is currently not available. |
663a60c55d | session | No description available. |
98bac1f50f | session | No description available. |
ac09458e72 | session | No description |
c72887300d | session | No description available. |
cbtest | 1 year | Description is currently not available. |
COMPASS | 1 hour | No description |
debug | never | No description available. |
DEVICE_INFO | 5 months 27 days | No description |
e188bc05fe | session | No description available. |
fd6b13af5c | session | No description available. |
gclid | 1 month | No description |
iamcsr | session | No description available. |
ncbi_sid | 1 year | No description available. |
pfjscookies | 1 year | Description is currently not available. |
ps_payloadSeqId | 2 hours | No description available. |
SM | session | No description available. |
VISITOR_PRIVACY_METADATA | 5 months 27 days | Description is currently not available. |
woopspro_recently_viewed_products | 7 days | Description is currently not available. |
zabHMBucket | 1 year | No description available. |
zc_consent | 1 year | No description available. |
zc_cu | 1 year | No description available. |
zc_cu_exp | 1 year | No description available. |
zc_loc | session | No description available. |
zc_show | 1 year | No description available. |
zc_tp | 1 year | No description available. |
zc_vopt | 1 year | Description is currently not available. |
ZCAMPAIGN_CSRF_TOKEN | session | No description available. |
zft-sdc | 1 hour 2 minutes | No description |
zps-tgr-dts | 1 year | No description |
zpssr* | session | No description |
zsc6339725afa6243a997dc48e2eac3fe89 | 30 minutes | No description |
zsd* | 2 hours | No description |
zsrdbO8X4S | 2 hours | No description |