posts explore innovations in microscopy, high throughput data capture and immuno-oncology through deep literature reviews and systematic data management.

Battle of the Science Bands

Battle of the Science Bands

Battle of the science bands ... Molecular Groove and The Checkpoints

To prevent wreckage, do not look directly at laser with remaining eye

Dear readers, I was not able to find any youtube videos of The Director's - the (defunct?) NIH house band, that I believe included Harold Varmus, Francis Collins, and Richard Klausner (and others?).


I was inspired to find science bands music videos today by short videos during Mark McKown's sample automation talk at today's (Oct 11, 2016) PerkinElmer seminar(s) at MD Anderson Cancer Center -- lunch catered by Goode Co., -- good choice!


The event was pretty good, title and bullet point summary ... my brief comments:


Advanced Sequencing, Imaging and Detection Applications 

  • AlphaScreen ... missed most of the talk (was recovering from Steve Rosenberg's immunotherapy talk from 8-9am).
  • HCS ... currently 3plex, slow, features expensive water dispenser objective lens ... could do at least 10x better if they talk with me (can start by reading my Brilliant Violet linkedin blog).
  • NGS sample Prep ... no single cell RNAseq + TCRseq + barcoded dextramer seq. 
  • Bioo mircoRNAseq and qRNAseq (Bioo Scientific was acquired by PerkinElmer in August 2016)  ... still no single cell RNAseq + paired TCRseq + barcoded dextramer seq (though maybe don't need every single RNA ... maybe simpler to go: paired TCRseq + barcoded dextramers + 100 key mRNAs, + 100 key lncRNAs + 100 key microRNAs). 

The event flyer is:

As a Prelude, Solo Artist Willem Overwijk performing "Cite Me"

One of his original adaptations, singing as usual on the shoulders of giants



OK, now what you've been waiting for: my favorite two science bands -- if you have recommendations for more, please let me know. I also included at bottom a couple of additional music videos.


Molecular Groove - Perkin Elmer's house band

No Grow Blues



Discovery - 2011



Discovery - Battle of the Biotech Bands, Royale Nightclub, Boston, MA

As seen here at this promising event, sure to be remembers in the pipeline of base hits on the path to the grand slam performance at the nobel after party ceremony.  

As seen here at this promising event, sure to be remembers in the pipeline of base hits on the path to the grand slam performance at the nobel after party ceremony.  


Web site (last updated 2011) 


linkedin page for Molecular Groove, PerkinElmer's house band

119 members,  is this (broken social circle?)

The Checkpoints -- the SITC house band. 


This is also the University of Texas MD Anderson Cancer Center's house band (I did not find any videos of Ron, Lynda, Ethan and Helen having jammed with them, nor The Chancellor, Admiral Willian McRaven). Band members include Jim Allison and Patrick Hwu. 


The first featuring Willem Overwijk (a.k.a. Prof. Adjuvantagous)

T-cells are forever - featuring Willem Overwijk



Come on baby, touch my dendrite (dendritic cell) / ? - The Checkpoints at SITC 2012



That ring of fire (?) - The Checkpoints Performance at SITC 2012 - Video #1


My public safety annoucement: "do not look at laser with remaining eye".


The Checkpoints Performance at SITC 2012 - Video #2



The Checkpoints perform at SITC 2014 (looks like Patrick Hwu on keyboards, but no Jim/harmonica)



The Checkpoints Perform at SITC's Fundraiser at the House of Blues Chicago


fun with ...


Molecules Gone Wild (Bio Style)" - Macromolecules Song (Music Video ver. Dance)



Before you go all hypoglycemic.


Glucose Glucose - Song by Science Groove



FYI - there are fluorescent protein biosensors (FPBs) for glucose, ATP, lactate and more. I just put together a table of FPBs for our update of a Current Protocols chapter on light microscopy and image analysis. Here is the 'direct from Excel' version:


Analyte or ActivityBiosensor(s)

  • 2-oxoglutarate (alpha-ketoglutarate) OGSor-GA, mOGsor, PROBS-2
  • ABL, BCR-ABL activityAbl indicator,  Picchu-Z/EGFR-Z, Prickle
  • activation of plasma membrane Ca2+ pumpBFP-PMCA-GFP
  • Akt (PKB)Eevee-iAkt, Aktus, BKAR, AktAR, ReAktion, GFP–Akt–YFP, GFP-PKB-RFP
  • AMPK activityAMPKAR, ABKAR, organelle specific ABKARs, T2AMPKAR (FLIM based)
  • Ammonium transport & concentrationAmTrac, MepTrac
  • Annexin A4 self-associationCYNEX4
  • arabinoseFLIParaF.Ec-250n
  • argininecpFLIPR, FPIPR-AhrC (CFP[-ahrC-YFP), FLIP-cpArgT194, FLIP-cpArtJ185, FRET arginine reporter
  • AutophagymTagRFP-LC3
  • ATPcyto-ATeam, GO-ATeam-2, ATeam3.10, AT1.03NL
  • ATP in mitochondriamit-Ateam (Kioka 2014)
  • ATP:ADP ratioPercevalHR (high dynamic range), Perceval
  • Aurora B kinaseaurora B sensor
  • BAI2 (a furanosyl borate diester)CLPY
  • BDNF concentration (extracellular)Becell (fusion protein TrkB ligand binding domain - EGFR kinase domain "BBD-Ecat", coexpressed with Ecaus biosensor) (on cell of interest or on "sentinel" cell). In principle, generalizable to any RTK.
  • Bile acid sensorsBAS
  • Ca2+ "GECIs": NTnC, YC Cameleons, NanoCaMeleons, GCaMP's, RCaMPs, D3cpV, TN-XXL, Twitch-1CD,  PeriCam's, Case12, Case16, Camgaroos, FIP-CBSM, FIP-CA3, FIP-CA9, FIP-CBSMTN-XL, B-GECO, R-GECO, jRCaMP1a, jRCaMP1b, Ca++ #2 (ECFP[]cp173Venus)
  • Ca2+ in endoplasmic reticulum (ER)D1ER, D4ER (low affinity Ca++ probes)
  • Ca2+ in lysosomesLAMP-1-YCaM
  • Ca2+ in mitochondria4mt-D3cpV
  • Ca2+ in vesiclesYcam2
  • Calcineurin activity (phosphatase)caNAR1
  • CaMKIICamuiα4m, CamuiαamCYCaMIIα
  • Caspase-3 protease activitySCAT3.1; CC3AI, RC3AI, VC3AI, mKate2-DEVD-iRFP, eqFP650-DEVD-iRFP
  • Caspase-3 protease activity, Caspase-6 activityCFP-c3-YFP-c6-mRFP
  • Caspase-6 protease activityCA6-GFP
  • Caspase-7 protease activityCA6-GFP
  • Caspase-8 protease activity (SCAT8.1 SCAT8.2
  • Cd2+Cd-FRET-2
  • Cdc42 activation Raichu-cdc42, A-probe.2
  • CDK/cyclin CDKsens, CDKact,  CyclinB1-CDK1 sensor
  • Cell cycle progression FUCCI, FUCCI2, PCNA-Chromobody,  HypoxCR
  • Cyclic AMP (cAMP)cADDis (cAMP Difference Detector in situ), Epac1-camps, Epac2-camps, RI-camps, ICUE, YR-ICUE,  HCN2-camps, (T)Epac(VV); FP-PKARII/FP-PKAcat, PKA Riα #7
  • Cyclic GMP (cGMP)Cygnet-1, Cygnet2, FlincG, CGYs, cGES, cGi, PKG #7 (cp173Venus-ECFP)
  • Citrate (citric acid)FLIPcit affinity series, CIT
  • Cl- CloMeleon,  LSSmClopHensor, Cl-sensor, EYFP (Chloride and other halide ions)
  • CREB activationICAP
  • Cu+Ace1-FRET, Mac1-FRET, Amt1-FRET
  • CyclinB-Cdk1CyclinB-Cdk1 sensor
  • Diacylglycerol (DAG)Upward DAG2, Upward DAG mNeon 1G8, Upward DAG mNeon 2D1, Downward DAG3, Daglas, DAGR, DIGDA, Cys-1-GFP, DAGR; CKAR (PKC activity)
  • EGFRFLAME, EGFR reporter, Ecaus, Bescell/ BBD-Ecat, EGFR-ECFP/PTV-EYFP
  • EGFR homodimerizationEGFR-VC & EGFR-VN (Bimolecular Fluorescence Complementation, BiFC; note: early versions irreversible)
  • EGFR interaction with Gαi3 (GTPase alpha subunit)EGFR-CFP & Gαi3-int YFP (internally tagged Galpha; also made "CT" C-terminal version)
  • EGFR–GIV–Gαi3 ternary complexesEGFR-VC & VN-GIV-CT (Bimolecular Fluorescence Complementation, BiFC) & Gαi3-int CFP (internally tagged; also made "CT" C-terminal version)
  • ERK (Extracellular signal-regulated kinase)EKAR, EKAR-EV, EKAR2G2, EAS, Erkus, Miu2, Erk KTR (kinase translocation reporter),  FIRE (prototype biosensor kinase activity by substrate stabilization)
  • Estrogen receptor (ER)CEY (ligand binding domain conformation)
  • FAK (Focal adhesion kinase)FAKbiosensor (Seong 2011)
  • Fyn (specific Src family kinase"Fyn sensor" (Yingiao Wang, UCSD; GM still waiting for "Lck this" sensor)
  • G protein activationFP-Gα/FP-Gβ(γ)
  • GlucokinaseFRET-GCK Reporters (mCerulean-GCK-mVenus)
  • Glucose FLIPglu affinity series, AcGFP1-GBPcys-mCherry, LoogerGlu
  • GlutamateiGluSnFr, SuperGluSnFR, GluSnFR, FLIPE affinity series, FLIP-cpGLtI210
  • glutamineFLIPQTV3.0 (affinity series) (glnH-mTFP1-glnH-Venus), FLIP-cpGlnH183, FRET glutamine reporter, LoogerGln
  • Granzyme BGrzB Reporter (Choi & Mictchison 2013), GrzB-Breakaway (McNamara, Korngold, Rabinovich propose CD19-TM-linker-mClover3-human optimized cleavage site-linker-mouse optimized cleavage site-linker-mRuby3-NoLS-mRuby3)
  • halide ionsCloMeleon, YFP-H148Q, YFP-H148QI152L, EYFP (quenched by chloride and other halide ions)
  • Hg2+eGFP205C, IFP/BV sensor
  • histidineFLIP-HisJ, FLIP-cpHisJ194
  • Histone H3 (K9) lysine methylationK9 reporter
  • Histone H3 (K27) lysine methylationK27 reporter
  • Histone H3 (S28) phosphorylation H3 phosphorylation reporter
  • Histone H4 lysine acetylationHistac-K5, Histac-K7, Histac-K12
  • inositol trisphosphate (IP3)LIBRA, FIRE1, FIRE-2, FIRE-3, IRIS-1
  • Insulin Receptor kinase activityPhocus-2pp, phocus, sinphos
  • JNK activityJuDAS, JNKAR; JNK KTR, controls: JNK KTR_AA (nuclear only), JNK KTR_EE (cytoplasm only)
  • LactateLaconic
  • Leucine FLIPLeu, FLIP-Leu-Y, FLIP-LivJ, FLIP-cpLivJ261
  • LysineFLIPK (affinity series)
  • M(1) muscarinic receptor activationM1R-YFP-CFP
  • Macromolecular crowdingCerulean-(GSG)6A(EAAAK)6A(GSG)6A(EAAAK)6A(GSG)6-Citrine
  • Malonyl-CoAFapR-D2eGFP
  • MaltoseFLIPmal affinity FRET series, Mal-### single FP series, EcMBP165-cpGFP.PPYF.T203V (Nadler 2016)
  • Mechanical straincpstFRET, stFRET, PriSSM, VinTS
  • MethionineFLIPM (Mohsin et al 2015, Mohsin and Ahmad 2014)
  • Mg2+Cerulean-HsCen3-Citrine
  • MT1-MMP activityMT1-MMP biosensor
  • Neurotrophic factorBBD-ECat/ECaus ("sentinel" cell-based)
  • Nitric oxide (NO) Piccell, FRET-MT, geNOp's (C-geNOp and M-geNOp excite at 430 nm, G-geNOp and Y-geNOp at 480 nm, O-geNOp at 515 nm; based on super enhanced cyan, mint green, EGFP, Venus yellow, mKOk)
  • O2 (Oxygen)FluBO, HIF-GFP,  8xHRE promoter-FP, HypoxCR,  dUnGOHR
  • O-GlcNAc transferaseO-GlcNAc sensor
  • Organic hydroperoxidesOHSer
  • p38 (MAPK)p38 KTR
  • PARP1 localization (poly(ADP-ribose) polymerase 1)PARP1 chromobody (nanobody-FP, recognizing ZnF2 domain, including aa 161, 188, 189) (Buchfellner 2016)
  • Pb2+ (lead ions)PbGFP (bright unbound, quenched by uM Pb2+ ions)
  • PCNA localization (proliferating cell nuclear antigen)PARP1 chromobody (nanobody-FP) (Burgess et al 2012)
  • PDK1 activation GFP-PDK1-RFP, PARE
  • pHSuper-ecliptic pHluorin, pHluorins (ecliptic, ratiometric), AlpHi, pHRed, SypHer, cpYFP, LSSmClopHensor, E2GFP, deGFP1-4, GFpH, YFpH, CFP-YFP tandem, pHuji
  • Phosphatidic acid (PA)Pii
  • Phosphatidylserine (PS)GFP-Lact-C2
  • PO4 (phsophate ion)FLIPPi
  • Protease activityMany: see Caspases, Granzyme B, for examples. iProtease, iTEV, iCasper (To 2015)
  • Protein kinase A (PKA) Epac’s, AKAR4, AKAR3, AKAR2, AKAR, AKAR1-34, ART, CRY-AKAR; PKA KTR
  • Protein kinase C (PKC)CKAR, δCKAR (PKC δ isoform specific), KCP-1
  • Protein kinase D (PKD)DKAR
  • protein dynamicsFast-FT (fast timer), Medium-FT (medium timer),  Slow-FT (slow timer); DsRed-Timer, many photochangable FPs
  • protein-protein interactionsmSplit reporters, using Bimolecular Fluorescence Complementation (BiFC) (generalizable)
  • Phosphoinositide’s dynamicsPIPline’s (Simon 2013)
  • PtdIns(3,4)P2Pippi-PI(3,4)P2; also binds PIP3: GFP-PHGRP1, GFP-PHPDK1, YFP-PHTAPP1, GFP-Svp1p, GFP-PHBtk, GFP-Ent3p, CAY
  • PtdIns(3,4,5)P3 (PIP3)Fllip, GFP-PHGRP1, GFP-Cytohesin-1, GFP-PHPDK1, YFP-PH-TAPP1, GFP-PH-Svp1p, GFP-PHBtk, GFP-PH,FP-CRAC, GFP-Ent3p, CAY, Pippi-PI(3,4,5)P3 … lipid binding domain (LBD) Pleckstrin homology domain derived from Grp1
  • PtdIns(4,5)P2Pippi-PI(4,5)P2, PPHPLCd1-YFP, GFP-ARNO, GFP-Cytohesin-1, GFP-PHOSH2, PHOSBP, GFP-PHFAPP, CAY
  • PtdIns(4,5)P2, Ins(1,4,5)P3 dynamicsGFP-PHPLC, FP-Tubby, FP-PHPLCδ1(FRET)
  • PtdIns(4,5)P2 / PLC activity CYPHER (PLC activity)
  • PtdIns(3)PGFP-FYVEEEA1, GFP–p40phox iPX, InPAkt
  • PutrescineFLIP-AF1
  • PyruvatePyronic (San Martin 2014)
  • QuinonesQsrY
  • Receptor tyrosine kinase activity (general) Picchu
  • redox and voltageroGFP, rxYFP149_202, CY-RL5, CY-RL7, HSP-FRET, Redoxfluor, HyPer, HyPEr2, mt-cpYFP, cpYFP
  • redox: H2O2HyPer, HyPer3, roGFP2-Orp1
  • redox: H2O2(picomolar H2O2)p130cas stabilized; PTEN oxidation blocks plasma membrane localization
  • redox: glutathioneroGFP2-Grx1
  • redox: superoxidemt-cpYFP, pcYFP
  • redox: PeroxiredoxinroGFP2-Prx
  • redox; thioredoxinTrx(CXXS)-cpYFP-Prx
  • redoxHSP-FRET
  • redox: NADH Frex, T-Rex,  rexYFP (rxYFP)
  • redox: NADH/NAD+ ratioPeredox, SoNAR
  • redox: NAD+"NAD+ Sensor" (Cambronne 2016)
  • redox: NADP+ Apollo-NADP+ (Cameron 2016)
  • redoxRedoxfluor
  • redox: RL5 RL7 (CY-RL7) ( Kolossov 2011)
  • Retinoic acidGEPRA
  • Rho family GTPase activation: cdc42Raichu-cdc42
  • Rho family GTPase activation: CRIBRaichu-CRIB
  • Rho family GTPase activation: Rac1Raichu-Rac1
  • Rho family GTPase activation: RalARaichu-Ral
  • Rho family GTPase activation: Rap1Raichu-Rap
  • Rho family GTPase activation: RasRaichu-Ras
  • Rho family GTPase activation: RhoARaichu-RhoA, RhoA2G
  • Rho family GTPase activation: RhoCRhoC FLARE
  • RhoGDI activityRaichu-RBD
  • RiboseFLIPrib affinity series
  • RTK–GIV–Gαi ternary complexesEGFR-VC & VN-GIV-CT (Bimolecular Fluorescence Complementation, BiFC) & Gαi3-int CFP (internally tagged) (generalized) (Midde et al 2015)
  • SERCA (Sarco/Endoplasmic Reticulum Ca(2+)-ATPase)CFP-SERCA
  • sphingosine-1-phosphateS1PR1-eGFP to S1PR1NB-tRFP (former is cell surface unless bound to S1P)
  • Src Srcus, BG-Src
  • SucroseFLIPsuc (affinity series)
  • synNOTCHSynthetic Notch signaling chimeric antigen receptors (CARs), i.e. extracellular binding domain-[key Notch activation components]-Gal4-VP64 artifical trancription factor (replacement for Notch ICD).
  • Taspase1 (Threonine Aspartase 1)TS-Cl2+ (NLS-GFP-GST-[MLL 2713KISQLDGVDD2722]-myc tag-NES (Multi-color translocation biosensor assays, when coexpressing Taspase1-BFP, cleavage results in NLS-GFP-GST localizing in nucleus) (see Granzyme B Breakaway for an alternative)
  • T-cell receptor signaling T-cell localization reporters (Singleton … Wulfing 2009, 2011 - series of 30 translocation reporters)
  • TrehaloseTre-C04 (Nadler 2016)
  • Trehalose-6-phosphateT6P-TRACK
  • TryptophanFLIPW-CTYT, FLIPW (affinity series)
  • Ubiquitin K63-linked sensorTAB2 NZF
  • Voltage membrane potential GEVI's: Arch3, eArch(D95N), Arclight, CaViar [Ca2+ and Vm], pHlaVor [pH and Vm], Mermaid,  Butterfly VSP3.1,  ASAP1,  Ace2N-2AA-mNeon,  MacQ-mCitrine; FlaSh, FlaSh-CFP/YFP, Flare, SPARC, VSFP-cpEGFP, VSFPs, PROPS
  • WASP (Wiskott -Aldrich syndrome protein ) conformationCdc42-GEF sensors
  • Zn2+eCALWY-6, eCALWY4, eZinCh, ZapCY1,  ZapCY2, ZifCY1, ZinCh, CLY9-2His, Cys2His2, GZnP1


Key References and Web Links

Newman, Fosbrink, Zhang 2011 Genetically encodable fluorescent biosensors for tracking signaling dynamics in living cells. Chem Rev 111: 3614-3666. (please see supplemental file).

Zhang, Ni, Newman (eds) (2014) Fluorescent Protein-Based Biosensors. Methods and Protocols. Methods in Molecular Biology 1071. Human Press. 251 pages. New York.

Addgene fluorescent protein guide: Biosensors




Many biosensors in this table use dim fluorescent proteins. EGFP is from 1996 and is less than 1/3rd the brightness of mNeonGreen or mClover3 (we hope that no reader is still driving a 1996 automobile or bicycle, unless it is a classic and you are showing off). This limits signal-to-noise ratio and dynamic range. We recommend replacement with bright FPs, such as mNeonGreen or mClover3 (green), mRuby3 (orange-red). Longer wavelengths (green, not blue or cyan) will also shift the fluorescence from high autofluorescence (NADH, NADPH, flavins, flavoproteins) to lower autofluorescence. We also recommend localizing the FP(s) if possible, for example, to plasma membrane (Lck-biosensor), or even to the channel/transporter being measured (i.e. GLUT4-linker-FLIPglu). We also suggest multimerizing FPs or FP biosensors whenever possible, for example LacI-nls-FP onto LacO 256x operator array on the same DNA plasmid (Robinett et al 1996). This could be made more efficeint now by replacing the original LacI-nls-EGFP with LacI-nls(mNeonGreen)3, which would be 10x brighter per protein molecule, enabling use of LacO 25x operator array. In the nucleus, one could localize to nucleolus (NoLS-mTFP1), PCNA replication factories (anti-PCNA nanobody-mNeonGreen), telomeres (Trf2-mRuby3), centromeres (CENP-mKate2), nuclear envelope(Nup-mCardinal2x2). See also Colorful Cell (Sladitschek and Neveu 2015), whose MXS-Chaining kit enables efficient cloning for multi-part expression (we do note that DNA plasmids enable two or more copies of similar or identical sequences, whereas manufacturing lentivirus or retroviruses typically results in deletions; this also has implications for expression of multiple shRNAs or microRNAs).


Sentinel cells: you could introduce an FP biosensor onto or into another cell. For example, to sense either adenosine or prostaglandin E2, you could introduce A2AR or EP2 G-protein coupled receptors (GPCRs), respectively, into HEK293 cells, along with a cyclic AMP biosensor. The biosensor could even be fused to the cell surface receptor. You could also knockout (CRISPR/Cas9 nuclease/nuckase, TALENs) or knockdown (RNAi by siRNA, shRNA or microRNA) all other receptors that signal through cyclic AMP. Optionally also KO/KD phosphodiesterases, though this could limit dynamic range; conversely, you could enforce expression of PDEs (maybe away from plasma membrane) to optimize both dynamic range and temporal sensitivity.

You could multiplex the sentinel cells by having different cell surface receptors (A2AR or EP2),  the same intracellular reporter (cAMP biosensor, and color code the Sentinel cells on either the receptor (mRuby3 vs mCardinalx2) or some out of the way location (i.e. nucleolus, with NoLS), or both (appropriate strength alternative splicing).

Translocation reporters, i.e. your favorite protein fused to a bright fluorescent protein ("fav"-mNeonGreen) are simple, powerful, tools. Singleton et al (2009, 2011) did this for 30 proteins in the T-cell receptor signaling pathway. Regot et al (2014) did this for four different kinase translocation reporters ("KTR"), including three simultaneously (JNK, p38 MAPK, Erk).

Histone marks can be reported on by FP fusion proteins to BET family members (BRDT, BRD2, and BRD4); histone acetyltransferases (TAFII250, PCAF, and GCN5), ATP dependent chromatin remodeling factors; or by nanobody-FP fusions (llama, camel, shark, mouse or human VHH antibody domains), scFv-FP fusions (single chain fragment variable), or by protein transfection of Fab-fluorophores ("FabLEM").

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