Many of our projects are chosen based on finding biological problems that can be studied in sufficient detail that we can do detailed biophysical modeling that can make predictions that we can test experimentally. Other projects involve developing new techniques that we believe will enable us to ask previously difficult to address questions. Some examples of each:
We are bringing a multidisciplinary approach to the study of a specific cancer, fibrolamellar hepatocellular carcinoma (FLC), a usually lethal liver cancer that affects adolescents and young adults Why this choice? It was based on the hopes:
- A well-defined cancer would be a homogeneous population, not a mixture of cancers.
- A cancer in adolescents would have a relatively clean background – the patients have not had decades of life to accumulate unrelated variations in the genome.
- A cancer in adolescents would be relatively “young” – the tumor has not had many decades to accumulate additional mutations.
If these hopes were correct, it would facilitate examining causes, progression, as well as developing diagnostics and therapeutics. This was part of the motivation behind the development of the Precision Medicine Initiative [we were honored to be invited by President Obama to introduce him, and the Precision Medicine initiative, at the launch at the White House and later to introduce Joe Biden and Pope Francis at the international launch of the Biden Cancer Initiative at The Vatican].
Validation of hopes:
We have shown that all fibrolamellar patients have a clean background with no common structural variants (inversions, amplifications, insertions/deletions) (Darcy, 2015) and only one common deletion: A loss of 400kb in one copy of chromosome 19 (Honeyman, 2014). This deletion results in the fusion gene of the first exon of DNAJB1, a heat shock protein co-factor, and the bulk of PRKACA, the catalytic subunit of protein kinase A. We have shown that the changes found in tumors, recurrences, metastases, in the coding transcriptome, the non-coding transcriptome, the proteome, the phosphoproteome, are almost indistinguishable between patients. We have shown that expression of this fusion protein, a chimera of DNAJB1-PRKACA is sufficient to produce the tumor in the livers of mice.
Working with patients:
We are actively engaged with the patient community, many of whom work with us at the bench.
- With the patients we have established the first tissue repository for fibrolamellar with tissue, blood, from over 140 patients)
- We work with the patients in establishing the first medical registry for fibrolamellar. This is owned and run by patients and their families. Over 200 patients have answered a collection of questions we have curated, many have uploaded their medical records, test and scans.
What have we learned?
We have shown that in all fibrolamellar patients there is a dysregulation of protein kinase A (Lalazar, 2018). In hundreds of patients, we have shown that the first exon of PRKACA, the catalytic subunit of protein kinase A, is replace with the first exon of DNAJB1, a heat shock protein co-factor (Honeyman, 2014). In one patient with fibrolamellar, we have found that they are missing a regulatory subunit of protein kinase A (Graham 2018). In three other patients the first exon of PRKACA is replaced with the first exon of ATP1B1, a sodium potassium ATPase. This dysregulation of protein kinase A produces changes in the coding and the non-coding transcriptome and the proteome that are:
- consistent from primary to metastases
- consistent between patients (Simon, 2015)
- distinct between fibrolamellar and other tumors (Simon, 2015)
Expression of a functional fusion oncokinase is sufficient to recapitulate this cancer:
- Using CRISPR in mouse liver to create the deletion, and form the fusion gene, is sufficient to produce the cancer.
- Expressing the fusion protein on its own, without any deletions, is sufficient to produce the cancer.
- A point mutation that eliminates the activity of the kinase, can no longer transform cells.
Where are we going?
What causes fibrolamellar?
- What leads to the break and fusion at that particular locus?
- We found that hundreds of patients who have a break in the first intron of DNAJB1 and the first intron of PRKACA. What is the structure of the DNA at the site of the break and fusion
- Why does the break not seen once people age?
- Why is it almost never seen outside the liver?
- The structural dynamics of the fusion protein (Cao et al., 2019; Tomasini et al., 2018)
- The impact of the fusion protein on the ecology of kinase signaling
- How does expression of the fusion oncokinase affect the cell biology of the liver cells? How does it:
- How do the altered liver cells interact with their environment and the immune system
- How do the structural dynamics of the fusion protein differ from the native kinase?
-Alter the transcriptome (Simon et al., 2015)
-Alter the proteome (Simon et al., 2015)
-Alter the phosphorylation landscape
Can we design ways of detecting the tumor and follow the tumor load in patients?
- We are developing these both to help detect recurrences earlier and to follow tumor load in patients in response to therapeutics
Can we develop therapeutics for fibrolamellar. Our strategies include:
- Targeting the fusion protein.
- Targeting the fusion transcript.
- Drug repurposing.
- Harnessing the immune system.
We are doing these studies in the following model systems:
- Patient-derived xenografts (Lalazar et al., 2021)
- Patient-derived organoids (Narayan et al., 2020; Saltsman et al., 2020)
- Genetically engineered mice (Kastenhuber et al., 2017)
- Cells freshly resected from patients
Below is a selection of our papers on fibrolamellar:
Cao B, Lu TW, Martinez Fiesco JA, Tomasini M, Fan L, Simon SM, Taylor SS, Zhang P. Structures of the PKA RIα Holoenzyme with the FLHCC Driver J-PKAcα or Wild-Type PKAcα. Structure. 2019 May 7;27(5):816-828.e4. doi: 10.1016/j.str.2019.03.001. Epub 2019 Mar 21. PubMed PMID: 30905674; PubMed Central PMCID: PMC6506387.
Dao Thi VL, Wu X, Belote RL, Andreo U, Takacs CN, Fernandez JP, Vale-Silva LA, Prallet S, Decker CC, Fu RM, Qu B, Uryu K, Molina H, Saeed M, Steinmann E, Urban S, Singaraja RR, Schneider WM, Simon SM, Rice CM. Stem cell-derived polarized hepatocytes. Nat Commun. 2020 Apr 3;11(1):1677. doi: 10.1038/s41467-020-15337-2.PMID: 32245952
Darcy DG, Chiaroni-Clarke R, Murphy JM, Honeyman JN, Bhanot U, LaQuaglia MP, Simon SM. The genomic landscape of fibrolamellar hepatocellular carcinoma: whole genome sequencing of ten patients. Oncotarget. 2015 Jan 20;6(2):755-70. doi: 10.18632/oncotarget.2712. PubMed PMID: 25605237; PubMed Central PMCID: PMC4359253.
Farber BA, Lalazar G, Simon EP, Hammond WJ, Requena D, Bhanot UK, La Quaglia MP, Simon SM. Non coding RNA analysis in fibrolamellar hepatocellular carcinoma. Oncotarget. 2018 Feb 13;9(12):10211-10227. doi: 10.18632/oncotarget.23325. eCollection 2018 Feb 13. PubMed PMID: 29535801; PubMed Central PMCID: PMC5828204.
Graham RP, Lackner C, Terracciano L, González-Cantú Y, Maleszewski JJ, Greipp PT, Simon SM, Torbenson MS. Fibrolamellar carcinoma in the Carney complex: PRKAR1A loss instead of the classic DNAJB1-PRKACA fusion. Hepatology. 2018 Oct;68(4):1441-1447. doi: 10.1002/hep.29719. Epub 2018 May 11. PubMed PMID: 29222914; PubMed Central PMCID: PMC6151295.
Hammond WJ, Lalazar G, Saltsman JA, Farber BA, Danzer E, Sherpa TC, Banda CD, Andolina JR, Karimi S, Brennan CW, Torbenson MS, La Quaglia MP, Simon SM. Intracranial metastasis in fibrolamellar hepatocellular carcinoma. Pediatr Blood Cancer. 2018 Apr;65(4). doi: 10.1002/pbc.26919. Epub 2017 Dec 29. PubMed PMID: 29286561; PubMed Central PMCID: PMC6028006.
Honeyman JN, Simon EP, Robine N, Chiaroni-Clarke R, Darcy DG, Lim II, Gleason CE, Murphy JM, Rosenberg BR, Teegan L, Takacs CN, Botero S, Belote R, Germer S, Emde AK, Vacic V, Bhanot U, LaQuaglia MP, Simon SM. Detection of a recurrent DNAJB1-PRKACA chimeric transcript in fibrolamellar hepatocellular carcinoma. Science. 2014 Feb 28;343(6174):1010-4. doi: 10.1126/science.1249484. PubMed PMID: 24578576; PubMed Central PMCID: PMC4286414.
Kastenhuber ER, Lalazar G, Houlihan SL, Tschaharganeh DF, Baslan T, Chen CC, Requena D, Tian S, Bosbach B, Wilkinson JE, Simon SM, Lowe SW. DNAJB1-PRKACA fusion kinase interacts with β-catenin and the liver regenerative response to drive fibrolamellar hepatocellular carcinoma. Proc Natl Acad Sci U S A. 2017 Dec 12;114(50):13076-13084. doi: 10.1073/pnas.1716483114. Epub 2017 Nov 21. PubMed PMID: 29162699; PubMed Central PMCID: PMC5740683.
Lalazar G, Requena D, Ramos-Espiritu L, Ng D, Bhola PD, de Jong YP, Wang R, Narayan NJC, Shebl B, Levin S, Michailidis E, Kabbani M, Vercauteren KOA, Hurley AM, Farber BA, Hammond WJ, Saltsman JA, Weinberg EM, Glickman JF, Lyons BA, Ellison J, Schadde E, Hertl M, Leiting JL, Truty MJ, Smoot RL, Tierney F, Kato T, Wendel HG, LaQuaglia MP, Rice CM, Letai A, Coffino P, Torbenson MS, Ortiz MV, Simon SM. Identification of Novel Therapeutic Targets for Fibrolamellar Carcinoma Using Patient Derived Xenografts and Direct from Patient Screening. Cancer Discov. 2021 Jun 14:candisc.0872.2020. doi: 10.1158/2159-8290.CD-20-0872. Online ahead of print. PMID: 34127480
Lalazar G, Simon SM. Fibrolamellar Carcinoma: Recent Advances and Unresolved Questions on the Molecular Mechanisms. Semin Liver Dis. 2018 Feb;38(1):51-59. doi: 10.1055/s-0037-1621710. Epub 2018 Feb 22. Review. PubMed PMID: 29471565; PubMed Central PMCID: PMC6020845.
Olivieri C, Walker C, Karamafrooz A, Wang Y, Manu VS, Porcelli F, Blumenthal DK, Thomas DD, Bernlohr DA, Simon SM, Taylor SS, Veglia G. Defective internal allosteric network imparts dysfunctional ATP/substrate-binding cooperativity in oncogenic chimera of protein kinase A. Commun Biol. 2021 Mar 10;4(1):321. doi: 10.1038/s42003-021-01819-6.PMID: 33692454 Free PMC article.
Rich BS, Honeyman JN, Darcy DG, Smith PT, Williams AR, Lim II, Johnson LK, Gönen M, Simon JS, LaQuaglia MP, Simon SM. Endogenous antibodies for tumor detection. Sci Rep. 2014 May 30;4:5088. doi: 10.1038/srep05088. PubMed PMID: 24875800; PubMed Central PMCID: PMC4038850.
Saltsman JA, Hammond WJ, Narayan NJC, Requena D, Gehart H, Lalazar G, LaQuaglia MP, Clevers H, Simon S.Cancers (Basel). A Human Organoid Model of Aggressive Hepatoblastoma for Disease Modeling and Drug Testing. 2020 Sep 18;12(9):2668. doi: 10.3390/cancers12092668.PMID: 32962010
Simon EP, Freije CA, Farber BA, Lalazar G, Darcy DG, Honeyman JN, Chiaroni-Clarke R, Dill BD, Molina H, Bhanot UK, La Quaglia MP, Rosenberg BR, Simon SM. Transcriptomic characterization of fibrolamellar hepatocellular carcinoma. Proc Natl Acad Sci U S A. 2015 Nov 3;112(44):E5916-25. doi: 10.1073/pnas.1424894112. Epub 2015 Oct 21. PubMed PMID: 26489647; PubMed Central PMCID: PMC4640752.
Simon JS, Botero S, Simon SM. Sequencing the peripheral blood B and T cell repertoire – Quantifying robustness and limitations. J Immunol Methods. 2018 Dec;463:137-147. doi: 10.1016/j.jim.2018.10.003. Epub 2018 Oct 10. PubMed PMID: 30312601; PubMed Central PMCID: PMC6355145.
Tomasini MD, Wang Y, Karamafrooz A, Li G, Beuming T, Gao J, Taylor SS, Veglia G, Simon SM. Conformational Landscape of the PRKACA-DNAJB1 Chimeric Kinase, the Driver for Fibrolamellar Hepatocellular Carcinoma. Sci Rep. 2018 Jan 15;8(1):720. doi: 10.1038/s41598-017-18956-w. PubMed PMID: 29335433; PubMed Central PMCID: PMC5768683.
- How do the different proteins fit together in an intact nuclear pore in a living cell?
- Is the structure static or dynamic?
We inserted a green fluorescent protein into many different positions of one of the structural components of the nuclear pore. We excited the GFP with polarized light. By varying the angle we could find the optimal angle for exciting the dipole of the GFP. With each amino acid we added in the alpha-helix at the GFP, the angle rotated 103o. We have the crystal structure of the GFP, thus, we could determine the orientation of the protein to which it was attached. We applied similar approach to the filaments that fill the lumen of the nuclear pore, allowing us to follow their dynamics. Next we applied the approach to various core structural domains and found some domains that changed with orientation with transport through the pore. The results were quantitative enough to allow us to make computational simulations of the structural dynamics and transport of the nuclear pore that we could then experimentally test. Some examples of our work:
- Mapping the orientation of nuclear pore proteins in living cells with polarized fluorescence microscopy.
- Nat Struct Mol Biol. 2011 Jun;18(6):643-9. doi: 10.1038/nsmb.2056. Epub 2011 Apr 17. PubMed PMID: 21499242; PubMed Central PMCID: PMC3109191.
- Kampmann M, Atkinson CE, Mattheyses AL, Simon SM.
- Conformation of the nuclear pore in living cells is modulated by transport state.
- 2020 Dec 21;9. doi: 10.7554/eLife.60654. PubMed PMID: 33346731; PubMed Central PMCID: PMC7752133.
- Pulupa J, Prior H, Johnson DS, Simon SM.
- Conserved spatial organization of FG domains in the nuclear pore complex.
- Biophys J. 2013 Jan 8;104(1):37-50. doi: 10.1016/j.bpj.2012.11.3823. Epub 2013 Jan 8. PubMed PMID: 23332057; PubMed Central PMCID: PMC3540264.
- Atkinson CE, Mattheyses AL, Kampmann M, Simon SM.
- Fluorescence anisotropy reveals order and disorder of protein domains in the nuclear pore complex.
- Biophys J. 2010 Sep 22;99(6):1706-17. doi: 10.1016/j.bpj.2010.06.075. PubMed PMID: 20858414; PubMed Central PMCID: PMC2941012.
- Mattheyses AL, Kampmann M, Atkinson CE, Simon SM.
- Simulations of nuclear pore transport yield mechanistic insights and quantitative predictions.
- Proc Natl Acad Sci U S A. 2011 Aug 2;108(31):E351-8. doi: 10.1073/pnas.1104521108. Epub 2011 Jun 20. PubMed PMID: 21690354; PubMed Central PMCID: PMC3150947.
- Mincer JS, Simon SM.
- A coarse-grained computational model of the nuclear pore complex predicts Phe-Gly nucleoporin dynamics.
- J Gen Physiol. 2017 Oct 2;149(10):951-966. doi: 10.1085/jgp.201711769. Epub 2017 Sep 8. PubMed PMID: 28887410; PubMed Central PMCID: PMC5694938.
- Pulupa J, Rachh M, Tomasini MD, Mincer JS, Simon SM.
We are using assembly of a virus as a model system for how a multimolecular machine assembles. Our approach has been to study single viruses as they assemble in a living cell. We study when and how individual viral encoded molecules and molecules in the host cell interact to allow the assembly of a functional virus. Most studies examine thousands, to millions of viruses in many cells, all in different states of infection, with viruses in different states of assembly. From examining single viruses, we found that the steps of assembly are highly co-ordinated with a very prescribed order of events. First, the genome comes to the surface, then the coat protein, Gag, assembles around it. As the Gag assembles, the membrane bends, folding around the nascent virion. Then a host protein, a member of the ESCRTIII complex, is recruited, followed within 5-10 seconds by another host protein, Vps4, an ATPase which remodels the ESCRT complex. They remain at the next of the nascent virion for under a minute, leave, and 20 seconds later the virus scissions from the host cell. The virus makes one main protein, Gag. It has a protease, which is activated upon assembly, which then cleaves Gag into separate pieces that then assemble inside the virion to mature the virus. Most recently we examined why viruses such as HIV-1 and Ebola don’t infect some species. We found that new world monkeys has a duplicated and truncate form one of the proteins of the ESCRTIII complex proteins. As result the normal ESCRT-III gets recruited, but fails to recruit the ATPase, so it falls away, but then comes back again a few times, until the ATPase is recruited and scission occurs. This produces a short few minute delay. However, it is enough for the virus to start activating its protease and cleave the Gag protein, which is then lost back into the cell, rather than being retained the virus. This few second delay does not seriously impact other ESCRT-III functions, such as cell division, but deleterious for the assembling virus. This suggests new potential therapeutic approaches to blocking the assembly of viruses such as HIV-1 and Ebola. A few examples of papers are:
- Imaging the biogenesis of individual HIV-1 virions in live cells.
- 2008 Jul 10;454(7201):236-40. doi: 10.1038/nature06998. Epub 2008 May 25. PubMed PMID: 18500329; PubMed Central PMCID: PMC2708942
- Jouvenet, Bieniasz, PD, Simon, SM.
- RetroCHMP3 blocks budding of enveloped viruses without blocking cytokinesis.
- 2021 Sep 27:S0092-8674(21)01055-2. doi: 10.1016/j.cell.2021.09.008. Online ahead of print.PMID:34597582
- Rheinemann L, Downhour DM, Bredbenner K, Mercenne G, Davenport KA, Schmitt PT, Necessary CR, McCullough J, Schmitt AP, Simon SM, Sundquist WI, Elde NC.
- Imaging the interaction of HIV-1 genomes and Gag during assembly of individual viral particles.
- Proc Natl Acad Sci U S A. 2009 Nov 10;106(45):19114-9. doi: 10.1073/pnas.0907364106. Epub 2009 Oct 27. PubMed PMID: 19861549; PubMed Central PMCID: PMC2776408.
- Jouvenet N, Simon SM, Bieniasz PD.
- Dynamics of ESCRT protein recruitment during retroviral assembly.
- Nature Cell Biology 2011 Apr;13(4):394-401. doi: 10.1038/ncb2207. Epub 2011 Mar 10. PubMed PMID: 21394083; PubMed Central PMCID: PMC3245320.
- Jouvenet N, Zhadina M, Bieniasz PD, Simon SM.
- Temporal and spatial organization of ESCRT protein recruitment during HIV-1 budding.
- Proc Natl Acad Sci U S A. 2014 Aug 19;111(33):12211-6. doi: 10.1073/pnas.1321655111. Epub 2014 Aug 6. PubMed PMID: 25099357; PubMed Central PMCID: PMC4142993
- Bleck, MF, Itano,MS, Johnson, DS, Thomas,VK, North,AJ, Bieniasz,PD, Simon,SM.
- Timing of ESCRT-III protein recruitment and membrane scission during HIV-1 assembly.
- 2018 Jul 4;7. doi: 10.7554/eLife.36221. PubMed PMID: 29972351; PubMed Central PMCID: PMC6080951.
- Johnson DS, Bleck M, Simon SM.
- Viral houseguests undertake interior redesign.
- 2010 May 28;141(5):754-6. doi: 10.1016/j.cell.2010.05.012. PubMed PMID: 20510923.
- Jouvenet N, Simon SM.
- Differential Regulation of Lipoprotein and Hepatitis C Virus Secretion by Rab1b.
- Cell Rep. 2017 Oct 10;21(2):431-441. doi: 10.1016/j.celrep.2017.09.053. PubMed PMID: 29020629; PubMed Central PMCID: PMC5685655.
- Takacs CN, Andreo U, Dao Thi VL, Wu X, Gleason CE, Itano MS, Spitz-Becker GS, Belote RL, Hedin BR, Scull MA, Rice CM, Simon SM.
- Readily Accessible Multiplane Microscopy: 3D Tracking the HIV-1 Genome in Living Cells.
- 2016 Feb;17(2):179-86. doi: 10.1111/tra.12347. Epub 2015 Dec 14. PubMed PMID: 26567131; PubMed Central PMCID: PMC4729623.
- Itano MS, Bleck M, Johnson DS, Simon SM.
- Recruitment of 7SL RNA to assembling HIV-1 virus-like particles.
- 2018 Jan;19(1):36-43. doi: 10.1111/tra.12536. Epub 2017 Dec 1. PubMed PMID: 29044909; PubMed Central PMCID: PMC6781622.
- Itano MS, Arnion H, Wolin SL, Simon SM.
- Plasma membrane is the site of productive HIV-1 particle assembly.
- PLoS Biol. 2006 Dec;4(12):e435. doi: 10.1371/journal.pbio.0040435. PubMed PMID: 17147474; PubMed Central PMCID: PMC1
- Jouvenet N, Neil SJ, Bess C, Johnson MC, Virgen CA, Simon SM, Bieniasz PD.
- And soon, a new paper from Kate Bredbenner in the lab (just accepted in Cell).
Check out the videos
- Ca2+ transients in melanocyte dendrites and dendritic spine-like structures evoked by cell-to-cell signaling.
- Journal of Cell Biology 2020 Jan 6;219(1). doi: 10.1083/jcb.201902014. PubMed PMID: 31821412.
- Belote RL, Simon SM.
Our studies with Gunter Blobel demonstrated that proteins moved across membranes through aqueous channels (see past projects). This forced the question: “What is the mechanism that moved the proteins”. This is part of a general question of how do molecular motors work. For movement of proteins across the membrane we proposed a biophysical mechanism which linked the chemical potential energy of translation to the movement of proteins, and we coined the term “Brownian Ratchet” (Simon et al. 1992) . The movement of the protein is dominated by Brownian (thermal) forces). The directionality was determined by various gradients of chemical potential in the cell. We have since worked on extrapolating this model to movement of molecules through the nuclear pore (Mincer and Simon, 2011).
How do other molecular motors function. The family of AAA+ ATPases are found in many cellular processes (we studied one member, VPS4, in our studies on viral assembly). Now we are examining ClpX, a component of a bacterial degradative machinery, akin to a proteasome. We are studying it both biochemically, but also we are pursuing single molecule studies of its activity using total internal fluorescence microscopy.
New results will be appearing here!
The immunological response in cancer:
- Simon JS, Botero S, Simon SM. Sequencing the peripheral blood B and T cell repertoire – Quantifying robustness and limitations. J Immunol Methods. 2018 Dec;463:137-147. doi: 10.1016/j.jim.2018.10.003. Epub 2018 Oct 10. PubMed PMID: 30312601; PubMed Central PMCID: PMC6355145.
- Rich BS, Honeyman JN, Darcy DG, Smith PT, Williams AR, Lim II, Johnson LK, Gönen M, Simon JS, LaQuaglia MP, Simon SM. Endogenous antibodies for tumor detection. Sci Rep. 2014 May 30;4:5088. doi: 10.1038/srep05088. PubMed PMID: 24875800; PubMed Central PMCID: PMC4038850.
This is a technique to facilitate our studies on cancer:
- Dao Thi VL, Wu X, Belote RL, Andreo U, Takacs CN, Fernandez JP, Vale-Silva LA, Prallet S, Decker CC, Fu RM, Qu B, Uryu K, Molina H, Saeed M, Steinmann E, Urban S, Singaraja RR, Schneider WM, Simon SM, Rice CM. Stem cell-derived polarized hepatocytes. Nat Commun. 2020 Apr 3;11(1):1677. doi: 10.1038/s41467-020-15337-2.PMID: 32245952
- Goulian M, Simon SM. Tracking single proteins within cells. Biophys J. 2000 Oct;79(4):2188-98. doi: 10.1016/S0006-3495(00)76467-8. PubMed PMID: 11023923; PubMed Central PMCID: PMC1301109.
- Itano MS, Bleck M, Johnson DS, Simon SM. Readily Accessible Multiplane Microscopy: 3D Tracking the HIV-1 Genome in Living Cells. Traffic. 2016 Feb;17(2):179-86. doi: 10.1111/tra.12347. Epub 2015 Dec 14. PubMed PMID: 26567131; PubMed Central PMCID: PMC4729623.
- Stabley DR, Oh T, Simon SM, Mattheyses AL, Salaita K. Real-time fluorescence imaging with 20 nm axial resolution. Nat Commun. 2015 Sep 22;6:8307. doi: 10.1038/ncomms9307. PubMed PMID: 26392382; PubMed Central PMCID: PMC4595625.
- Jaiswal JK, Mattoussi H, Mauro JM, Simon SM. Long-term multiple color imaging of live cells using quantum dot bioconjugates. Nat Biotechnol. 2003 Jan;21(1):47-51. doi: 10.1038/nbt767. Epub 2002 Dec 2. PubMed PMID: 12459736.
- Jaiswal JK, Goldman ER, Mattoussi H, Simon SM. Use of quantum dots for live cell imaging. Nat Methods. 2004 Oct;1(1):73-8. doi: 10.1038/nmeth1004-73. PubMed PMID: 16138413.
- Voura EB, Jaiswal JK, Mattoussi H, Simon SM. Tracking metastatic tumor cell extravasation with quantum dot nanocrystals and fluorescence emission-scanning microscopy. Nat Med. 2004 Sep;10(9):993-8. doi: 10.1038/nm1096. Epub 2004 Aug 29. PubMed PMID: 15334072.
- Jaiswal JK, Simon SM. Potentials and pitfalls of fluorescent quantum dots for biological imaging. Trends Cell Biol. 2004 Sep;14(9):497-504. doi: 10.1016/j.tcb.2004.07.012. Review. PubMed PMID: 15350978.
- Koeppel F, Jaiswal JK, Simon SM. Quantum dot-based sensor for improved detection of apoptotic cells. Nanomedicine (Lond). 2007 Feb;2(1):71-8. doi: 10.2217/174358184.108.40.206. PubMed PMID: 17716192.
- Jaiswal JK, Simon SM. Imaging Live Cells Using Quantum Dots. Cold Spring Harb Protoc. 2015 Jul 1;2015(7):619-25. doi: 10.1101/pdb.top086322. PubMed PMID: 26134913.
Some of the techniques trying to advance the use of total internal reflection:
- Schmoranzer J, Goulian M, Axelrod D, Simon SM. Imaging constitutive exocytosis with total internal reflection fluorescence microscopy. J Cell Biol. 2000 Apr 3;149(1):23-32. doi: 10.1083/jcb.149.1.23. PubMed PMID: 10747084; PubMed Central PMCID: PMC2175105.
- Johnson DS, Toledo-Crow R, Mattheyses AL, Simon SM. Polarization-controlled TIRFM with focal drift and spatial field intensity correction. Biophys J. 2014 Mar 4;106(5):1008-19. doi: 10.1016/j.bpj.2013.12.043. PubMed PMID: 24606926; PubMed Central PMCID: PMC4026788.
- Johnson DS, Jaiswal JK, Simon S. Total internal reflection fluorescence (TIRF) microscopy illuminator for improved imaging of cell surface events. Curr Protoc Cytom. 2012 Jul;Chapter 12:Unit 12.29. doi: 10.1002/0471142956.cy1229s61. PubMed PMID: 22752951; NIHMSID
- Mattheyses AL, Simon SM, Rappoport JZ. Imaging with total internal reflection fluorescence microscopy for the cell biologist. J Cell Sci. 2010 Nov 1;123(Pt 21):3621-8. doi: 10.1242/jcs.056218. Review. PubMed PMID: 20971701; PubMed Central PMCID: PMC2964103.
- Fix M, Melia TJ, Jaiswal JK, Rappoport JZ, You D, Söllner TH, Rothman JE, Simon SM. Imaging single membrane fusion events mediated by SNARE proteins. Proc Natl Acad Sci U S A. 2004 May 11;101(19):7311-6. doi: 10.1073/pnas.0401779101. Epub 2004 May 3. PubMed PMID: 15123811; PubMed Central PMCID: PMC409915.
- Jaiswal JK, Simon SM. Total internal reflection fluorescence microscopy for high-resolution imaging of cell-surface events. Curr Protoc Cell Biol. 2003 Nov;Chapter 4:Unit 4.12. doi: 10.1002/0471143030.cb0412s20. PubMed PMID: 18228434.