DMMD0203_MD023L_spec.(LEV专用-中英文版)_20151015.pdf

P.O. BOX 44994 •MADISON WI 53744.4994 •PHONE: +1.608.441.2323 •TOLL FREE: 800.700.2282GRASON-STADLER NICOLET BIOMEDICAL NICOLET VASCULAR TOENNIESPRODUCT SPECIFICATIONS November 2004 | GSI TympStar | 2000-0112 Rev1The following specifications apply to both Version 1and Version 2 of the GSI TympStar. > Bold arrowed type indicates the additional capabilities of Version 2.GENERAL SPECIFICATIONS W x D x H: 52 cm x 38 cm x 32 cm Weight: 7.5 Kg Shipping Weight: 13.4 Kg Power Consumption: 120 Watts maximum Test Types: Tympanometry, Acoustic Reflex Threshold,Reflex Decay, Eustachian T ube Function (Intact & Perforated )Protocols: Diagnostic, Screening, User-defined > Special Tests: Two-component T ympanometry,Multiple Frequency T ympanometry, Acoustic Reflex Latency Test, Reflex Sensitization Display: Internal or External VGA Monitor Interface: RS232, parallel and keyboard output Printout: Internal or External Deskjet or Laserjet Printer PROBE TONE 226 Hz (85 dB SPL ±1.5 dB)> 678 Hz (85 dB SPL ±3.0 dB)> 1000 Hz (75 dB SPL ±3.0 dB)Accuracy: ±1%Harmonic Distortion: Less than 5%ADMITTANCE MEASUREMENTS Range: 226 Hz (-1.0 to +7.0)> 678 Hz (-5.0 to +25)> 1000 Hz (-5.0 to +30)Sensitivity Scale: Auto Scales to Appropriate Range,Manual selection also possible in Reflex Modes only Accuracy (226 Hz): Tymp Mode: ±5% of reading or ±0.1 ml, whichever is greater Reflex Mode: ±5% of reading or ±0.2 ml, whichever is greater PRESSURE MEASUREMENTS (load volume of 0.2 to 7.0 ml)Range:Normal = +200 to -400 daPa Wide = +400 to -600 daPa Accuracy: ±10% of reading or ±10 daPa, whichever is greater Sweep Rate: 12.5, 50.0 and 600/200 daPa/sec, > 200 daPa/sec.Sweep Accuracy: 10% of nominal rate Maximum limits (in 0.5cc cavity ): -800 daPa & +600 daPa REFLEX MEASUREMENTS Stimuli: > 250, 500, 1k, 2k, 4k, BBN, LBN, HBN,> Click (100 microseconds pulse), External Input,Non-acoustic Frequency Accuracy: ±3%Harmonic Distortion (THD): Less than 5% (measured acoustically)Noise Signals: (3 dB bandwidths)Low Band: 125 -1,600 Hz High Band: 1,600 -4,000 Hz Broad Band: 125 -4,000 Hz Intensity Range: 35 to 120 dB HLStep Size: 5 dB, >1 dB and 2 dB Calibration Accuracy: ±3 dB Step Accuracy: ±0.5 dB ON/OFF Ratio: 70 dB minimum ENVIRONMENTAL Temperature:Storage: -40°C to +75°C Operating: +15°C to +35°C Humidity: 90% at 35°C (non-condensing)ACCESSORIES SUPPLIED Probe assembly (including contralateral insert phone)Eartips (1 pkg. each standard, special, screening)Printer paper, 2 rolls thermal, 1 roll self-adhesive Calibration test cavity, Cleaning kit, Probe mount kit (shoulder, clip, wrist band), Quick User Guide,Reference Instruction Manual, Remote (RS232 link)Dust CoverOPTIONAL ACCESSORIES Deskjet printer 1700-9613External VGA monitor 1700-9614External keyboard 1700-9615Isolation Transformer 1700-9617Service Manual 2000-0110Conversion Kit 2000-9650(V1 to V2)LANGUAGE KITS GSI TympStar Version 1English 2000-9645E French 2000-9645F German 2000-9645G Spanish 2000-9645S Italian 2000-9645I GSI TympStar Version 2English 2000-9646E French 2000-9646F German 2000-9646G Spanish 2000-9646S Italian 2000-9646I QUALITY SYSTEM Manufactured, designed, developed and marketed by VIASYS Healthcare Inc. NeuroCare Group under ISO 13485, ISO 9001 certified quality system.COMPLIANCE / REGULATORY STANDARDSDesigned, tested and manufactured to meet the following domestic (USA), Canadian, European and International Standards:UL 2601-1American Standards for Medical Electrical Equipment. IEC 601-1, EN 60601-1International Standards for Medical Electrical Equipment.CSA C22.2 # 601-1-M90Medical Device Directive (MDD) (ID No.: 0344) to comply with “EC Directive” 93/42/EECANSI S3.39, ANSI S3.6, IEC 645-1, IEC 1027, ISO 389GSI TYMPSTAR ™Middle Ear Analyzer。

B0300ug Rev 04/231User GuideBetaine Solution5 M, PCR ReagentB0300Product DescriptionBetaine, also called trimethylglycine orN,N,N-triethylammonium acetate, is an analog of glycine with three methyl groups.1 Betaine is a PCR enhancing reagent that is widely used for improving the yield and specificity of PCR products, especially for the amplification of targets rich in GC content or those that form secondary structures resulting in poor yield. Betaine facilitates DNA strand separation and decreases secondary structure of GC-rich regions.2 Betaine has been broadly used to optimize multiplex and ‘long and accurate′ polymerase chain reaction (LA-PCR). The addition of 1.0-1.7 M aqueous betaine to a PCR mixture has been reported to reduce the base pair composition dependence on DNA strand melting.3Quality Specifications•Suitable for molecular biology applications with high GC content. Upon addition of BetaineSolution to 1.2 M, a 994 bp human gene target with 64% GC content is amplified via PCR with JumpStart™ Taq DNA Polymerase, while no amplification is detected in the absence of Betaine Solution.•DNase free: No degradation of HindIII-digested lambda phage DNA detected after incubation with 1.2 M Betaine Solution for 16 hours at 37 °C. •RNase free: No degradation of tRNA detected after incubation with 1.2 M Betaine Solution for 16 hours at 37 °C.•Endonuclease free: No nicking or linearization of pBR322 plasmid DNA detected after incubation with 1.2 M Betaine Solution for 16 hours at 37 °C. Applications• Loop-mediated isothermal amplification (LAMP)4 • PCR for genomic DNA amplification 5 •Quantitative PCR (qPCR)6 and RT-qPCR• PCR amplification of CGG repeats in genomic DNA 7•Reverse transcription for single cell cDNA library preparation via Smart-seq-total 8Intended UseThis product is for R&D use only. Not for drug,household, or other uses. Please consult the Safety Data Sheet for information regarding hazards and safe handling practices.StorageStore at 2-8 °C.Directions for UseAdd Betaine to a final concentration of 1.0 – 1.7 M in a nucleic acid amplification mixture prior to initiation of thermocycling/heating.References1. Day CR and Kempton SA. Biochim biophys acta.,1860(6):1098-106 (2016). 2. Jensen MA, et al. PLoS ONE, 5(6):e11024.(2010). 3. Rees WA, et al. Biochem., 32(1):137-44. (1993). 4. Kostic T, et al. Appl Microbiol Biotechnol.,99(18): 7711–7722. (2015). 5. Azaiez H, et al. Hum Mutat., 24(4):305-11.(2004). 6. Milte CM, et al. Eur J Nutr., 57(1):363-372.(2018). 7. Saulto A, et al. J Mol Diagn., 7(5):605-12.(2005). 8. Isakova A, et al. Proc Natl Acad Sci USA.,118(51):e2113568118. (2021).The life science business of Merck operatesas MilliporeSigma in the U.S. and Canada.Merck and Sigma-Aldrich are trademarks of Merck KGaA, Darmstadt, Germany or its affiliates.All other trademarks are the property of their respective owners. Detailed information on trademarks is available via publicly accessible resources.© 2023 Merck KGaA, Darmstadt, Germany and/or its affiliates. All Rights Reserved.B0300ug Rev 04/232Product OrderingOrder products online at .Description Catalogue NumberJumpStart™ Taq DNAPolymerase D4184Deoxynucleotide (dNTP)Mix, containing 10 mMeach of dATP, dCTP, dGTP,and dTTP sodium saltsD7295 Nuclease-free water W1754Custom ordered primersspecific to gene target OLIGO GenElute™-E Single SpinDNA Cleanup Kit EC600 GenElute™ PCRClean-Up Kit NA1020 GenElute™ GelExtraction Kit NA1111Precast Agarose Gels P6222 P5472 P6097 P5972 P57221 kb DNA Ladder D0428 Water, Microbial DNA-free MBD0025 Nuclease-Free Water,for Molecular Biology W4502 JumpStart™ Taq Ready Mix P2893 RED Taq® Ready Mix P0982 Glycerol-free JumpStart™Taq DNA Polymerase D9310 DMSO D8418 Single strandbinding protein S3917 BST Max DNA Polymerase SRE0113 NoticeWe provide information and advice to our customers on application technologies and regulatory matters to the best of our knowledge and ability, but without obligation or liability. Existing laws and regulations are to be observed in all cases by our customers. This also applies in respect to any rights of third parties. Our information and advice do not relieve our customers of their own responsibility for checking the suitability of our products for the envisaged purpose. The information in this document is subject to change without notice and should not be construed as a commitment by the manufacturing or selling entity, or an affiliate. We assume no responsibility for any errors that may appear in this document. Technical AssistanceVisit the tech service page at/techservice.Terms and Conditions of SaleWarranty, use restrictions, and other conditions of sale may be found at /terms. Contact InformationFor the location of the office nearest you, go to /offices.。

1.IDENTIFICATION OF THE SUBSTRANCE/PREPARATION AND THECOMPANY/UNDERTAKINGPOSITION/INFORMATION ON INGREDIENTS3.HAZARDS IDENTIFICATIONMaterial Safety Data SheetRevision Date:February 23,2018Product code LT005,LT006Product name Lenti-Pac™HIV qRT-PCR Titration KitsContact manufacturer GeneCopoeia,Inc.9620Medical Center Drive,Suite 101Rockville,MD 20850USAPhone:301-762-0888Toll free:1-866-360-9531Fax:301-762-3888Chemical Name CAS-No Weight %Phenol108-95-230-60Guanidine isothiocyanate 593-84-015-40Ammonium thiocyanate 1762-95-47-13Glycerol56-81-540-70Contact with acids or bleach liberates toxic gases.DO NOT ADD acids or bleach to any liquid wastes containing this product.We recommend handling all chemicals with caution.GHS –Classification Signal Word DANGERHealth HazardsAcute oral toxicity Category 4Acute dermal toxicityCategory 4Acute Inhalation Toxicity -Vapors Category 4Skin corrosion/irritationCategory 1B Serious eye damage/eye irritationCategory 1Specific target organ systemic toxicity (single exposure)Category 3Specific target organ systemic toxicity(repeated exposure)Category3Mutagenicity Mutagenic category2Physical hazardsNot hazardousHazard StatementsH314-Causes severe skin burns and eye damageH341-Suspected of causing genetic defectsH373-May cause damage to organs through prolonged or repeated exposureH412-Harmful to aquatic life with long lasting effectsH302-Harmful if swallowedH312-Harmful in contact with skinH332-Harmful if inhaledH335-May cause respiratory irritationPrecautionary StatementsP301+P312-IF SWALLOWED:Call a POISON CENTER or doctor/physician if you feel unwellP304+P340-IF INHALED:Remove victim to fresh air and keep at rest in a position comfortable for breathingP301+P330+P331-IF SWALLOWED:rinse mouth.Do NOT induce vomitingP303+P361+P353-IF ON SKIN(or hair):Remove/Take off immediately all contaminated clothing.Rinse skin withwater/showerP305+P351+P338-IF IN EYES:Rinse cautiously with water for several minutes.Remove contact lenses,if present and easy to do.Continue rinsingP310-Immediately call a POISON CENTER or doctor/physicianP308+P313-IF exposed or concerned:Get medical advice/attentionP273-Avoid release to the environmentP280-Wear protective gloves/protective clothing/eye protection/face protectionPrinciple Routes of ExposurePotential Health EffectsEyes Causes burns.Risk of serious damage to eyes.Corrosive to the eyesand may cause severe damage including blindness.Skin Causes burns.Possible risk of irreversible effects.Harmful in contactwith skin.Irritating to skin and mucous membranes.Inhalation Harmful by inhalation.Ingestion Harmful if swallowed.Ingestion causes burns of the upper digestive andrespiratory tracts.Ingestion may cause gastrointestinal irritation,nausea,vomiting and diarrhea.Specific effectsCarcinogenic effects Phenol has been classified by the International Agency for Research onCancer(IARC)as not classifiable as to carcinogenicity to humans(Group3).Mutagenic effects Not ApplicableReproductive toxicity Not ApplicableSensitization Not ApplicableTarget Organ Effects SkinLungsLiverSpleenKidney4.FIRST AID MEASURES5.FIRE-FIGHTING MEASURES6.ACCIDENTAL RELEASE MEASURES7.HANDLING AND STORAGEHMISHealth 3*Chronic HazardFlammability 1ReactivitySkin contact Wash off immediately with soap and plenty of water while removing all contaminated clothes and shoes.Call a physician immediately.Eye contactIF IN EYES:Rinse cautiously with water for several minutes.Remove contact lenses,if present and easy to do.Continue rinsing.Rinseimmediately with plenty of water,also under the eyelids,for at least 15minutes.Call a physician immediately.Ingestion Call a physician or poison control center immediately.Rinse mouth.Do not induce vomiting without medical advice.Never give anything by mouth to an unconscious person.InhalationRemove to fresh air.Call a physician or poison control center immediately.Notes to physicianTreat symptomatically.Suitable extinguishing media Dry chemical.Carbon dioxide (CO2).Water spray.Foam.Special protective equipment for firefightersWear self-contained breathing apparatus and protective suit.Australia HazChem Code2XPersonal precautionsELIMINATE all ignition sources (no smoking,flares,sparks or flames in immediate area).Use personal protection equipment.Avoid contact with skin,eyes or clothing.Ensure adequate ventilation.Keep people away from and upwind of spill/leak.Evacuate personnel to safe areas.Methods for cleaning upPrevent product from entering drains.Soak up with inert absorbentmaterial.Neutralize spill with slaked lime,sodium bicarbonate or crushed limestone.Collect powdered material and deposit in sealed containers and dispose of phenol as hazardous waste.Isolate area and deny entry.Environmental precautionsPrevent further leakage or spillage if safe to do so.Prevent product from entering drains.Do not allow material to contaminate ground water system.See Section 12for more information8.EXPOSURE CONTROLS /PERSONAL PROTECTIONHandling Always wear recommended Personal Protective Equipment.Avoid contact with skin,eyes or clothing.Remove all sources of ignition.StorageKeep containers tightly closed in a cool,well-ventilated place.Keep away from heat,sparks,flame and other sources of ignition (i.e.,pilot lights,electric motors and static electricity).Protect from sunlight.Exposure limitsChemical name OSHA PELOSHA PEL (Ceiling)ACGIH OEL (TWA)ACGIH OEL (STEL)Phenol5ppm 19mg/m 3None 5ppm None Guanidine isothiocyanate None None None None Ammonium thiocyanate 5mg/m 3None None None Glycerol15mg/m 3None10mg/m 3NoneEngineering measures Use in a chemical fume hoodPersonal protective equipmentPersonal Protective Equipment requirements are dependent on the user institution's risk assessment and are specific to the risk assessment for each laboratory where this material may be used.Respiratory protection In case of insufficient ventilation wear suitable respiratory equipment RespiratorUp to 50ppmRecommendations,National Institute of Occupational Safety and Health,U.S.(APF =10)Any air-purifying half-mask respirator with organic vaporcartridge(s)in combination with an N95,R95,or P95filter.The following filters may also be used:N99,R99,P99,N100,R100,P100.(APF =10)Any supplied-air respirator Up to 125ppm:(APF =25)Any supplied-air respirator operated in a continuous-flow mode.(APF =25)Any powered air-purifying respirator with an organic vapor cartridge in combination with a high-efficiency particulate filter.Up to 250ppm:(APF =50)Any air-purifying full-facepiece respirator equipped with organic vapor cartridge(s)in combination with an N100,R100,or P100filter.(APF =50)Any air-purifying,full-facepiece respirator (gas mask)with a chin-style,front-or back-mounted organic vapor canister having an N100,R100,or P100filter.(APF =50)Any powered,air-purifying respirator with a tight-fitting facepiece and organic vapor cartridge(s)in combination with a high-efficiency particulate filter.(APF =50)Any self-contained breathing apparatus with a full facepiece.(APF =50)Any supplied-air respirator with a full facepiece.Emergency or planned entry into unknown concentrations or IDLH conditions:(APF =10,000)Any self-contained breathing apparatus that has a full facepiece and is operated in a pressure-demand or other positive-pressure mode.9.PHYSICAL AND CHEMICAL PROPERTIES10.STABILITY AND REACTIVITY11.TOXICOLOGICAL INFORMATION(APF =10,000)Any supplied-air respirator that has a full facepiece and is operated in a pressure-demand or other positive-pressure mode in combination with an auxiliary self-contained positive-pressure breathing apparatus.Escape:(APF =50)Any air-purifying,full-facepiece respirator (gas mask)with a chin-style,front-or back-mounted organic vapor canister having an N100,R100,or P100filter./Any appropriate escape-type,self-contained breathing apparatus.Hand protectionImpervious gloves.S24-Avoid contact with skin.S36-Wear suitable protective clothing.Eye protectionTight sealing safety goggles.Skin and body protection Impervious clothing.Hygiene measuresContaminated work clothing should not be allowed out of the workplace.Avoid contact with skin,eyes or clothing.Handle in accordance with good industrial hygiene and safety practice.Environmental exposure ControlsPrevent product from entering drains.General information FormLiquid.Appearance Red,maroon.OdorMedicinal,sweet,tar-like.Boiling point/range °C No data available °F No data available Melting point/range °C No data available °F No data available Flash point°C No data available °F No data available Autoignition temperature °C No data available °F No data availableOxidizing properties No information available Water solubilitySolubleStabilityStable under normal conditions.Materials to avoidStrong oxidizing agents.Strong acids.Isocyanates.Heat.Nitriles,Nitrides.Alkaline earth metals.Strong oxidizers,alkali metals and alkaline earth metals may cause fires or explosions.Hazardous decomposition Toxic gas.Sulphur oxides.Hydrogen cyanide (hydrocyanic acid).Carbon oxides,ProductsNitrogen Oxides.PolymerizationHazardous polymerization does not occur.Acute toxicityChemical name LD50(oral,rat/mouse)LD50(dermal,rat/rabbit)LC50(inhalation,rat/mouse)Phenol=317mg/kg (Rat)No data available =316mg/m3(Rat)Guanidine isothiocyanate571mg/kg2000mg/kg5.319mg/L (4H)12.ECOLOGICAL INFORMATIONAmmonium thiocyanate =500mg/kg (Rat)No data available No data available Glycerol =12600mg/kg (Rat)No data available No data availablePrinciple Routes of Exposure Potential Health Effects Eyes Causes burns.Risk of serious damage to eyes Corrosive to the eyes and may cause severe damage including blindness.Skin Causes burns.Possible risk of irreversible effects Harmful in contact with skin.Irritating to skin and mucous membranes.Inhalation Harmful by inhalationIngestionHarmful if swallowed Ingestion causes burns of the upper digestive and respiratory tracts Ingestion may cause gastrointestinal irritation,nausea,vomiting and diarrheaCarcinogenic effects Phenol has been classified by the International Agency for Research on Cancer (IARC)as not classifiable as to carcinogenicity to humans (Group 3).Mutagenic effects No information available.Reproductive toxicity No information available.SensitizationNo information available.Target organ effectsSkin.Lungs.Liver.Spleen.Kidney.EcotoxicityHarmful to aquatic organisms,may cause long-term adverse effects in the aquatic environment.Chronic aquatic toxicity Category 3MobilitySee log PowBiodegradation Inherently biodegradable Bioaccumulation No information availableChemical name Freshwater algae data Water flea data Freshwater fish species dataMicrotox dataLog Pow PhenolDesmodesmusDaphnia magna =316mg/m3(Rat)logPow1.subspicatus EC50187EC5047-279mg/L (72h) 4.24-10.7mg/L Pseudokirchneriella (48subcapitata EC50h)Daphnia 46.42mg/L (96h)magnaEC5010.2-15.5mg/L (48h)13.DISPOSAL CONSIDERATIONS14.TRANSPORT INFORMATION15.REGULATARY INFORMATIONGlycerolDaphnia magna EC50>500mg/L (24h)logPow1.76Dispose of contents/containers in accordance with local regulations.IATAProper shipping name Corrosive liquid,n.o.s.(guanidine thiocyanate-phenol solution).Hazard Class 8Subsidiary class None Packing group II UN-No 1760ERG Code153ComponentTSCA Phenol,108-95-2(30-60)Listed Guanidine isothiocyanate,593-84-0(15-40)Listed Ammonium thiocyanate 1762-95-4(7-13)Listed Glycerol,56-81-5(40-70)ListedUS Federal Regulations SARA 313This product contains the following toxic chemical(s)subject to the notification requirements of section 313of Title III of the Superfund Amendments and Reauthorization Act of 1986.This law requires certain manufacturers to report on annual emissions of specified chemicals and chemical categories.Please note that if you repackage,or otherwise redistribute,this product to industrial customers,a notice similar to this one should be sent to those customers:Chemical name CAS-No.Weight %SARA 313-Threshold values Phenol108-95-230-60 1.0Ammonium thiocyanate1762-95-47-131.0Clean Air Act,Section 112Hazardous Air Pollutants (HAPs)(see 40CFR 61)This product contains the following HAPs:Component CAS-No.Weight %HAPS data Phenol108-95-230-60PresentAmmonium thiocyanate1762-95-47-13Present (XCN where X=H or any16.OTHER INFORMATIONother group where a formal dissociation may occur.For example KCN or Ca[CN]2)US state regulations Chemical name Massachusetts -RTK New Jersey-RTK Pennsylvania-RTK Illinois-RTK Rhode Island-RTK PhenolListed Listed Listed Listed Listed Guanidine isothiocyanate -----Ammonium thiocyanate Listed -Listed Listed Listed GlycerolListed-Listed-ListedCalifornia Proposition 65This product does not contain any Proposition 65chemicals.WHMIS Hazard Class D1A -Very toxic materials E -Corrosive materialThis product has been classified in accordance with the hazard criteria of the Controlled Products Regulations (CPR)and the MSDS contains all the information required by the CPR.Reason for revision Not Applicable.SDS sections updated.For research use only."The above information was acquired by diligent search and/or investigation and the recommendations are based on prudent application of professional judgment.The information shall not be taken as being all inclusive and is to be used only as a guide.All materials and mixtures may present unknown hazards and should be used withcaution.Since the Company cannot control the actual methods,volumes,or conditions of use,the Company shall not be held liable for any damages or losses resulting from the handling or from contact with the product as described herein.THE INFORMATION IN THIS SDS DOES NOT CONSTITUTE A WARRENTY,EXPRESSED OR IMPLIED,INCLUDING ANY IMPLIED WARRANTY OF MERCHANTABILITY OR FITNESS FOR ANY PARTICULAR PUPOSE"End of Safety Data Sheet。

AlphaScreen®SureFire®STAT3 (p-Tyr705) Assay KitsManualAssay Points Catalog #500TGRS3S50010 000TGRS3S10K50 000TGRS3S50KFor Research Use OnlyResearch Reagents for Research Purposes OnlyGeneral Information on the AlphaScreen® SureFire® p-STAT3 assayThe AlphaScreen® SureFire® p-STAT3 assay is used to measure the phosphorylation of endogenous STAT3αand STAT3βin cellular lysates. The assay is an ideal system for the screening of both modulators of receptor activation (e.g. agonists and antagonists) as well as agents acting intracellularly, such as small molecule inhibitors of upstream events. The assay will measure STAT3 phosphorylation by either cloned or endogenous receptors, and can be applied to primary cells.This assay eliminates the need for laborious techniques, such as Western blotting or conventional ELISA. It is a homogeneous assay, in that no sample washing steps are required, which allows for minimal handling, short assay times, and robotic operation if desired. The assay utilizes the bead-based Alpha Technology, and requires an Alpha Technology-compatible plate reader.Alpha Technology AlphaScreen® SureFire® Assay PrincipleAlphaScreen® SureFire® technology allows the detection of phosphorylated proteins in cellular lysates in a highly sensitive, quantitative and user friendly assay. In these assays, sandwich antibody complexes, which are only formed in the presence of analyte, are captured by AlphaScreen donor and acceptor beads, bringing them into close proximity. The excitation of the donor bead provokes the release of singlet oxygen molecules that triggers a cascade of energy transfer in the Acceptor beads, resulting in the emission of light at 520-620nm.Kit-Specificity informationThis assay kit contains antibodies which recognize the phospho-Tyr705-epitope, and a distal epitope on STAT3αand STAT3β. The proteins detected by this kit correspond to GenBank Accessions NP_644805 and NP_998827.These antibodies recognize STAT3 of human, mouse and rat origin. Other species should be tested on a case-by-case basis.Kit ContentsKit Size500 points10,000 points50,000 pointsLysis buffer (5X) 1 x 10 mL 4 x 60 mL 3 x 400 mL Activation buffer 1 x 2 mL 1 x 60 mL 1 x 300 mL Reaction buffer 1 x 2.6 mL 1 x 45 mL 1 x 225 mL Dilution buffer 1 x 1.5 mL1 x 25 mL2 x 60 mLPositive Control Lysate 1 tube to be re-dissolved in 250 µL H 2O Negative Control Lysate1 tube to be re-dissolved in 250 µL H 2OStorage conditions upon receiptThe kit buffers e.g. 5X Lysis buffer, Activation buffer and Reaction buffer should be stored at 4°C. DO NOT freeze the kit buffers – the Reaction buffer contains antibodies and freeze/thaw cycles can lead to a loss of activity.Materials Required But Not ProvidedThe AlphaScreen SureFire assay kits are optimized to work with AlphaScreen Protein A general IgG detection beads. These are available separately from PerkinElmer. The AlphaScreen Protein A general IgG detection kits contain a biotinylated rabbit IgG control, which can be used to test the instrument settings and bead performance.ItemSuggested source Catalog # Size Protein A general IgG detection kit(contains the Acceptor and Donor Beads) PerkinElmer Inc. 6760617C 6760617M 6760617R 500 pt 10,000 pt 50,000 pt Proxiplate™-384 Plus, white, shallow well assay platePerkinElmer Inc. 6008280 6008289 50/box 200/box Optiplate™-384 Plus, white, assay plate PerkinElmer Inc. 6007290 6007299 50/box 200/box TopSeal-A 384, clear adhesive sealing film PerkinElmer Inc. 6050185100/boxEnvision® or Enspire® Alpha-readerPerkinElmer Inc.--Buffer preparation and subsequent storage conditions5X Lysis bufferStore 5X Lysis buffer at 4°C. For assay, dilute 5-fold in water immediately prior to use. Discard unused buffer. Activation bufferPrecipitation will occur during storage 4°C. To re-dissolve, warm to 37°C and mix. Alternatively, Activation buffer can be stored at room temperature with no loss in activity.Reaction buffer*Keep on ice while in use. Do not freeze.Once diluted discard unused reaction buffer. AlphaScreen ® Protein A IgG KitStore at 4°C in the dark.Acceptor Mix(Reaction buffer + Activation buffer +AlphaScreen ® Acceptor beads)Immediately prior to use, dilute Activation buffer B 5-fold in Reaction buffer (e.g. take 98 μL Activation buffer and dilute in 392 μL Reaction buffer).Dilute Acceptor beads 50-fold in Acceptor mix (e.g. add10 μL Acceptor beads to 490 μL of premixed Reactionbuffer + Activation buffer).The Acceptor mix should be used immediately for best results. Excess mix should be discarded.Donor Mix**(Dilution buffer + AlphaScreen ®Donor beads)Immediately prior to use, dilute Donor beads 20-fold in Dilution buffer (e.g. add 10 μL Don or beads to 190 μLDilution buffer).The Donor mix should be used immediately for best results. Excess mix should be discarded.Assay Control lysateAfter reconstitution in 250 μL water, lysates should be frozen at -20°C in single use aliquots and used within 1 month.* Do not vortex the Reaction buffer, as vigorous mixing can damage some antibodies. ** Prepare and use Donor Mix under low-light conditions.Control Lysate informationControl lysates are prepared from A431 cells (ATCC #CRL-1555) at a concentration of approximately 1 mg/mL. The controls are supplied lyophilized, and should be reconstituted in either dd H 2O or MilliQ® H 2O. Once reconstituted, lysates should be stored frozen in single use aliquots.Negative Lysate: Prepared from A431 cells treated with EGF receptor inhibitor (2 μM AG1478)for 2 hours prior to lysis.Positive Lysate: Prepared from A431 cells treated with 200 ng/mL EGF for 10 minutes.p-STAT3 AlphaScreen® SureFire® Assay ProtocolsA. 2-Plate Assay - assay protocol for adherent cellsCell Seeding1. Seed cell s (200 μL of cells for 96 well plates, 50 μL for 384 well plates) in tissue culture plates. Incubate at 37°C overnight in serum-containing media.Cell Treatment2. Remove culture media, and stimulate the cells with 50 μL agonists prepared in serum-free media (25 µL for 384-well plates). (If testing antagonists, prior to stimulation remove culture medium and replace with 50 μL serum-free media containing antagonists (25 µL for 384-well plates)). Return cells to 37°C incubator for desired time. 1 hour is often sufficient for signal transduction inhibitors, and 5 minutes for receptor agonists.Note: Peptidic agonists and antagonists can often stick to plastic surfaces. To minimize this effect, dilute in serum-free media containing a suitable carrier protein (e.g. 0.1% IgG free BSA - Jackson Immunoresearch Cat #001-000-161).Lysate Preparation5. To lyse cells, remove medium from wells, and add freshly prepared 1X Lysis Buffer(50-100 μL fora 96 well plate, 25 μL for a 384 well plate). Agitate on a plate shaker (~350 rpm) for 10 minutes at room temperature.6. Take 4 μL of the lysate and transfer to a 384-well Proxiplate™ for assay. (Add 4 μL Control lysates to separate wells if required).SureFire Assay7. Add 5 μL of Acceptor Mix to wells. Seal plate with Topseal-A adhesive film, and incubate for 2 hours at room temperature.8. Add 2 μL of Donor Mix to wells under subdued light. Seal plate with Topseal-A adhesive film, and cover plate with foil. Incubate for 2 hours at room temperature.Note: Longer incubation may give greater sensitivity. Plates can be incubated overnight if required.9. Read plate on an Alpha Technology-compatible plate reader, using standard AlphaScreen settings.B. 1 Plate Assay - assay protocol for non-adherent cells, and for high-throughput applications. Note: the larger volumes required using this assay will result in achieving less assay points per kit. Cell Seeding1. Harvest cells by centrifugation, and re-suspend cells in HBSS at a suitable cell density. We recommend 107cells/mL as a starting point. Seed 4 μL of cells/well into a 384-well culture plate.2. If using test agents/inhibitors, add 2 μL/well of 4X inhibitors prepared in HBSS.Note: Peptidic agonists and antagonists can often stick to plastic surfaces. To minimize this effect, dilute in serum-free media containing a suitable carrier protein (e.g. 0.1% IgG free BSA - Jackson Immunoresearch Cat #001-000-161).3. Return cells to incubator at 37°C for 1-2 hours.Cell Treatment4. Stimulate cells with agonists by addition of 2 μL/well of 4X agonist stock in HBSS containing 0.1% BSA. The final volume in the wells should be 8 μL. (if no antagonists were used in step 2, stimulate the cells with 4 μL/well of 2X agonist, to give a final volume in the wells of 8 μL.)Lysate Preparation5. To lyse the cells, add 2 μL/well 5X Lysis buffer.(Add 10 μL control lysates to separate wells if required)SureFire Assay6. Add 8 μL of Acceptor Mix to wells. Seal plate with Topseal-A adhesive film, and incubate for 2 hours at room temperature.7. Add 3 μL of Donor Mix to wells under subdued light. Seal plate with Topseal-A adhesive film, and cover plate with foil. Incubate for 2 hours at room temperature.Note: Longer incubation may give greater sensitivity. Plates can be incubated overnight if required.8. Read plate on an Alpha Technology-compatible plate reader, using standard AlphaScreen settings.Representative DataWestern blot analysis (35 μg protein/lane) of phospho-STAT3 in lysates generated from either unstimulated (-) or IL6-stimulated (+) THP-1 cells, or the AlphaScreen SureFire assay (3.5 μg protein/well), using the standard 2-plate protocol.THP-1 cells (ATCC Cat# TIB-202) were harvested from growing flasks, resuspended in HBSS, and seeded at 25K cells/well in 384-well microplates. Cells were incubated for 2 hours at 37°C, and stimulated with IL-6 for 10 minutes at room temperature (left) or pre-treated with JAK inhibitor I for 2 hours at 37°C, then stimulated with IL-6 (right). Cells were lysed with 5X Lysis buffer with shaking at RT for 10 minutes, and analyzed for phospho-STAT3 using the standard 1-plate protocol.Frequently Asked Questions & TroubleshootingFor comprehensive information on assay optimization and troubleshooting, please refer to the following resources:▪ Guide to AlphaScreen® SureFire ® assay optimization ▪ AlphaScreen® SureFire ® user guideTo download these resources, and other related technical information, visit /category/alpha-surefire-kitsFor general information on AlphaScreen® SureFire ® assays, visit - +Customer CareTo contact the customer care team, please visit /ServiceCallFor more information regarding related AlphaScreen® SureFire® products andprotocols refer to:PerkinElmer web site: TGR BioSciences website: FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.This product is not for resale or distribution except by authorized distributors.LIMITED WARRANTY: PerkinElmer, Inc. warrants that, at the time of shipment, the products sold by it are free from defects in material and workmanship and conform to specifications which accompany the product. PerkinElmer Inc. makes no other warranty, express or implied with respect to the products, including any warranty of merchantability or fitness for any particular purpose. Notification of any breach of warranty must be made within 60 days of receipt unless provided in writing by PerkinElmer Inc. No claim shall be honored if the customer fails to notify PerkinElmer Inc. within the period specified. The sole and exclusive remedy of the customer for any liability of PerkinElmer Inc. of any kind including liability based upon warranty (express or implied whether contained herein or elsewhere), strict liability contract or otherwise is limited to the replacement of the goods or the refunds of the invoice price of goods. PerkinElmer Inc. shall not in any case be liable for special, incidental or consequential damages of any kind.。