UMBC Institute of Fluorescence (IoF)

Permanent URI for this collectionhttp://hdl.handle.net/11603/10906

The IoF was founded in 2001 by Dr. Chris D. Geddes, Professor and Director. Under the leadership of Dr. Chris D. Geddes, the IoF has earned a well-deserved international reputation for its advances in Fluorescence Spectroscopy and Plasmonics. Approaches and concepts both developed and discovered by the group, such as Metal-Enhanced Fluorescence (MEF), Metal-Enhanced Chemiluminescence (MEC), Surface-Plasmon Coupled Phenomenon and the glucose-sensing contact lens, are both well-recognized, highly cited and currently used in laboratories around the world today.

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Fixed-angle observation of surface plasmon coupled chemiluminescence from palladium thin films
(AIP, 2009-09-25) Aslan, Kadir; Weisenberg, Micah; Hortle, Elinor; Geddes, Chris
In this letter, the fixed-angle observation of surface plasmon coupled chemiluminescence (SPCC) from palladium thin films is reported. Fresnel calculations predict that light 492–549 nm spectral range can efficiently induce surface plasmon modes in 15 nm palladium thin films. Free-space emission from blue, green, and chartreuse chemiluminescent solutions on palladium thin films was isotropic, while the SPCC emission was highly directional and predominantly p-polarized in accordance with the predictions of the Fresnel calculations. In addition, the decay rates of the SPCC and free-space emission were similar, which suggests that palladium thin films have no catalytic effect on chemiluminescence emission.
Indium nanodeposits: A substrate for metal-enhanced fluorescence in the ultraviolet spectral region
(AIP, 2010-11-03) Dragan, Anatoliy; Geddes, Chris
We have studied a metallic substrate, composed of indium nanodeposits, for metal-enhanced fluorescence (MEF) in the ultraviolet (UV) spectral region. Indium coated slides were prepared using the thermal vapor deposition technique. Theoretical finite difference time domain simulations and experimental studies show that plasmon enhanced absorption and coupled radiation through the scattering component of the extinction spectra of indium nanoparticles, lie in UV region, and are sensitive to the size and density of the nanoparticles, the thickness of the indium film, and polarity of the medium. The MEF effect, measured for intrinsic protein tryptophan and tyrosine residues, loaded onto indium films of different thickness, changes in a wavelike fashion, reflecting changes in the metal film landscape and, consequently, the chromophores coupling with surface plasmons. Indium films also significantly enhance intrinsic fluorescence of proteins themselves [bovine serum albumin]. In this case the wavelength dependence of MEF shows different emission enhancements of protein Tyr and Trp residues. Subsequently, indium-enhanced intrinsic protein fluorescence in the UV spectral region can be of great potential importance for quantitation assays as well as for the labeless detection of biomolecules in the biosciences.
Metal-enhanced fluorescence from tin nanostructured surfaces
(AIP, 2010-01-20) Zhang, Yongxia; Dragan, Anatoliy; Geddes, Chris
The recent surge in interest in the metal-enhanced fluorescence (MEF) phenomenon and its numerous applications in the biosciences has fueled research into identifying alternative metals to silver which have desirable properties, such as enhanced emission and fluorophore photostability. In this paper, we subsequently study and reveal that tin nanodeposits are a suitable metal for MEF with an electric field wavelength dependence somewhat different than silver. An enhanced fluorescence emission coupled with a reduced fluorophore lifetime suggests both an electric field and plasmon-coupling component are the underlying mechanisms for tin-based MEF. In addition, an enhanced fluorophore photostability is observed near-to tin nanodeposits.
Metal-enhanced chemiluminescence from chromium, copper, nickel, and zinc nanodeposits: Evidence for a second enhancement mechanism in metal-enhanced fluorescence
(AIP, 2010-09-28) Weisenberg, Micah; Zhang, Yongxia; Geddes, Chris
Over the past decade metal-fluorophore interactions, metal-enhanced fluorescence, have attracted significant research attention, with the technology now becoming common place in life science applications. In this paper, we address the underlying mechanisms of metal-enhanced fluorescence (MEF) and experimentally show using chemiluminescence solutions that MEF is indeed underpinned by two complimentary mechanisms, consistent with the recent reports by Geddes and co-workers [Zhang et al., J. Phys. Chem. C 113, 12095 (2009)] and their enhanced fluorescence hypothesis.
Development of a Microwave—Accelerated Metal-Enhanced Fluorescence 40 Second, <100 cfu/mL Point of Care Assay for the Detection of Chlamydia Trachomatis
(IEEE, 2011-03) Zhang, Yongxia; Agreda, Patricia; Kelley, Shannon; Gaydos, Charlotte; Geddes, Chris
An inexpensive technology to both lyse Chlamydia trachomatis (CT) and detect DNA released from CT within 40 s is demonstrated. In a microwave cavity, energy is highly focused using 100-nm gold films with “bow-tie” structures to lyse CT within 10 s. The ultrafast detection of the released DNA from less than 100 cfu/mL CT is accomplished in an additional 30 s by employing the microwave-accelerated metal-enhanced fluorescence technique. This new “ release and detect” platform technology is a highly attractive alternative method for the lysing of bacteria, DNA extraction, and the fast quantification of bacteria and potentially other pathogenic species and cells as well. Our approach is a significant step forward for the development of a point of care test for CT.
Ultra-Fast and Sensitive Detection of Non-Typhoidal Salmonella Using Microwave-Accelerated Metal-Enhanced Fluorescence (“MAMEF”)
(PLOS, 2011-04-08) Tennant, Sharon M.; Zhang, Yongxia; Galen, James E.; Geddes, Chris; Levine, Myron M.
Certain serovars of Salmonella enterica subsp. enterica cause invasive disease (e.g., enteric fever, bacteremia, septicemia, meningitis, etc.) in humans and constitute a global public health problem. A rapid, sensitive diagnostic test is needed to allow prompt initiation of therapy in individual patients and for measuring disease burden at the population level. An innovative and promising new rapid diagnostic technique is microwave-accelerated metal-enhanced fluorescence (MAMEF). We have adapted this assay platform to detect the chromosomal oriC locus common to all Salmonella enterica subsp. enterica serovars. We have shown efficient lysis of biologically relevant concentrations of Salmonella spp. suspended in bacteriological media using microwave-induced lysis. Following lysis and DNA release, as little as 1 CFU of Salmonella in 1 ml of medium can be detected in <30 seconds. Furthermore the assay is sensitive and specific: it can detect oriC from Salmonella serovars Typhi, Paratyphi A, Paratyphi B, Paratyphi C, Typhimurium, Enteritidis and Choleraesuis but does not detect Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Streptococcus pneumoniae, Haemophilus influenzae or Acinetobacter baumanii. We have also performed preliminary experiments using a synthetic Salmonella oriC oligonucleotide suspended in whole human blood and observed rapid detection when the sample was diluted 1:1 with PBS. These pre-clinical data encourage progress to the next step to detect Salmonella in blood (and other ordinarily sterile, clinically relevant body fluids).
Sialyl Residues Modulate LPS-Mediated Signaling through the Toll-Like Receptor 4 Complex
(PLOS, 2012-04-09) Feng, Chiguang; Stamatos, Nicholas M.; Dragan, Anatoliy; Medvedev, Andrei; Whitford, Melissa; Zhang, Lei; Song, Chang; Rallabhandi, Prasad; Cole, Leah; Nhu, Quan M.; Vogel, Stefanie N.; Geddes, Chris; Cross, Alan S.
We previously reported that neuraminidase (NA) pretreatment of human PBMCs markedly increased their cytokine response to lipopolysaccharide (LPS). To study the mechanisms by which this occurs, we transfected HEK293T cells with plasmids encoding TLR4, CD14, and MD2 (three components of the LPS receptor complex), as well as a NFₖB luciferase reporting system. Both TLR4 and MD2 encoded by the plasmids are α-2,6 sialylated. HEK293T cells transfected with TLR4/MD2/CD14 responded robustly to the addition of LPS; however, omission of the MD2 plasmid abrogated this response. Addition of culture supernatants from MD2 (sMD2)-transfected HEK293T cells, but not recombinant, non-glycosylated MD2 reconstituted this response. NA treatment of sMD2 enhanced the LPS response as did NA treatment of the TLR4/CD14-transfected cell supplemented with untreated sMD2, but optimal LPS-initiated responses were observed with NA-treated TLR4/CD14-transfected cells supplemented with NA-treated sMD2. We hypothesized that removal of negatively charged sialyl residues from glycans on the TLR4 complex would hasten the dimerization of TLR4 monomers required for signaling. Co-transfection of HEK293T cells with separate plasmids encoding either YFP- or FLAG-tagged TLR4, followed by treatment with NA and stimulation with LPS, led to an earlier and more robust time-dependent dimerization of TLR4 monomers on co-immunoprecipitation, compared to untreated cells. These findings were confirmed by fluorescence resonance energy transfer (FRET) analysis. Overexpression of human Neu1 increased LPS-initiated TLR4-mediated NFₖB activation and a NA inhibitor suppressed its activation. We conclude that (1) sialyl residues on TLR4 modulate LPS responsiveness, perhaps by facilitating clustering of the homodimers, and that (2) sialic acid, and perhaps other glycosyl species, regulate MD2 activity required for LPS-mediated signaling. We speculate that endogenous sialidase activity mobilized during cell activation may play a role in this regulation.
Metal-enhanced fluorescence: The role of quantum yield, Q, in enhanced fluorescence
(AIP, 2012-03-02) Dragan, Anatoliy; Geddes, Chris
Metal-enhanced fluorescence has attracted enormous research and commercial interest in recent years, due to the ability to significantly enhance fluorescence signatures in the near-field as well as protect fluorophores against photobleaching. In this article, we address one of the major unresolved questions, whether far-field fluorophore quantum yield, Q₀, has a direct relationship to fluorescence enhancement factors in metal-enhanced fluorescence.
Ultra-fast pg/ml anthrax toxin (protective antigen) detection assay based on microwave-accelerated metal-enhanced fluorescence
(Elsevier, 2012-06-01) Dragan, Anatoliy; Albrecht, Mark T.; Pavlovic, Radmila; Keane-Myers, Andrea M.; Geddes, Chris
Rapid presymptomatic diagnosis of Bacillus anthracis at early stages of infection plays a crucial role in prompt medical intervention to prevent rapid disease progression and accumulation of lethal levels of toxin. To detect low levels of the anthrax protective antigen (PA) exotoxin in biological fluids, we have developed a metal-enhanced fluorescence (MEF)–PA assay using a combination of the MEF effect and microwave-accelerated PA protein surface absorption. The assay is based on a modified version of our “rapid catch and signal” (RCS) technology previously designed for the ultra-fast and sensitive analysis of genomic DNA sequences. Technologically, the proposed MEF–PA assay uses standard 96-well plastic plates modified with silver island films (SiFs) grown within the wells. It is shown that the fluorescent probe, covalently attached to the secondary antibody, plays a crucial role of indicating complex formation (i.e., shows a strong MEF response to the recognition event). Microwave irradiation rapidly accelerates PA deposition onto the surface (“rapid catch”), significantly speeding up the MEF–PA assay and resulting in a total assay run time of less than 40min with an analytical sensitivity of less than 1pg/ml PA.
Blind Evaluation of the Microwave-Accelerated Metal-Enhanced Fluorescence Ultrarapid and Sensitive Chlamydia trachomatis Test by Use of Clinical Samples
(ASM, 2020-12-21) Melendez, Johan; Huppert, Jill S.; Jett-Goheen, Mary; Hesse, Elizabeth A.; Quinn, Nicole; Gaydos, Charlotte A.; Geddes, Chris
Accurate point-of-care (POC) diagnostic tests for Chlamydia trachomatis infection are urgently needed for the rapid treatment of patients. In a blind comparative study, we evaluated microwave-accelerated metal-enhanced fluorescence (MAMEF) assays for ultrafast and sensitive detection of C. trachomatis DNA from vaginal swabs. The results of two distinct MAMEF assays were compared to those of nucleic acid amplification tests (NAATs). The first assay targeted the C. trachomatis 16S rRNA gene, and the second assay targeted the C. trachomatis cryptic plasmid. Using pure C. trachomatis, the MAMEF assays detected as few as 10 inclusion-forming units/ml of C. trachomatis in less than 9 min, including DNA extraction and detection. A total of 257 dry vaginal swabs from 245 female adolescents aged 14 to 22 years were analyzed. Swabs were eluted with water, the solutions were lysed to release and to fragment genomic DNA, and MAMEF-based DNA detection was performed. The prevalence of C. trachomatis by NAATs was 17.5%. Of the 45 samples that were C. trachomatis positive and the 212 samples that were C. trachomatis negative by NAATs, 33/45 and 197/212 were correctly identified by the MAMEF assays if both assays were required to be positive (sensitivity, 73.3%; specificity, 92.9%). Using the plasmid-based assay alone, 37/45 C. trachomatis-positive and 197/212 C. trachomatis-negative samples were detected (sensitivity, 82.2%; specificity, 92.9%). Using the 16S rRNA assay alone, 34/45 C. trachomatis-positive and 197/212 C. trachomatis-negative samples were detected (sensitivity, 75.5%; specificity, 92.9%). The overall rates of agreement with NAAT results for the individual 16S rRNA and cryptic plasmid assays were 89.5% and 91.0%, respectively. Given the sensitivity, specificity, and rapid detection of the plasmid-based assay, the plasmid-based MAMEF assay appears to be suited for clinical POC testing.
Spectral shifts in metal-enhanced fluorescence
(AIP, 2014-08-11) Karolin, Jan; Geddes, Chris
We report a 2 nm red shift in the fluorescence spectra observed for Rhodamine 800 dissolved in glycerol on copper substrates as compared to glass reference samples, suggesting a wavelength dependence of metal enhanced fluorescence. The full width half maximum of the blue-red spectra is about 1 nm narrower as compared to the reference sample. We speculate that the observation correlates with a specific interaction mechanism between the Rhodamine 800 transition dipole, the enhanced electric field, and subsequent plasmon coupling, an observation not yet reported.
Adaptation of red blood cell lysis represents a fundamental breakthrough that improves the sensitivity of almonella detection in blood
(Wiley, 2015-01-29) Boyd, M. A.; Tennant, S. M.; Melendez, Johan; Toema, D.; Galen, J. E.; Geddes, Chris; Levine, M. M.
Aims Isolation of Salmonella Typhi from blood culture is the standard diagnostic for confirming typhoid fever but it is unavailable in many developing countries. We previously described a Microwave Accelerated Metal Enhanced Fluorescence (MAMEF)-based assay to detect Salmonella in medium. Attempts to detect Salmonella in blood were unsuccessful, presumably due to the interference of erythrocytes. The objective of this study was to evaluate various blood treatment methods that could be used prior to PCR, real-time PCR or MAMEF to increase sensitivity of detection of Salmonella. Methods and Results We tested ammonium chloride and erythrocyte lysis buffer, water, Lymphocyte Separation Medium, BD Vacutainer® CPT™ Tubes and dextran. Erythrocyte lysis buffer was the best isolation method as it is fast, inexpensive and works with either fresh or stored blood. The sensitivity of PCR- and real-time PCR detection of Salmonella in spiked blood was improved when whole blood was first lysed using erythrocyte lysis buffer prior to DNA extraction. Removal of erythrocytes and clotting factors also enabled reproducible lysis of Salmonella and fragmentation of DNA, which are necessary for MAMEF sensing. Conclusions Use of the erythrocyte lysis procedure prior to DNA extraction has enabled improved sensitivity of Salmonella detection by PCR and real-time PCR and has allowed lysis and fragmentation of Salmonella using microwave radiation (for future detection by MAMEF). Significance and Impact of the Study Adaptation of the blood lysis method represents a fundamental breakthrough that improves the sensitivity of DNA-based detection of Salmonella in blood.
National Institute of Biomedical Imaging and Bioengineering Point-of-Care Technology Research Network: Advancing Precision Medicine
(IEEE, 2016-08-16) Ford Carleton, Penny; Schachter, Steven; Parrish, John A.; Collins, John M.; Crocker, J. Benjamin; Dixon, Ronald F.; Edgman-Levitan, Susan; Lewandrowski, Kent B.; Stahl, James E.; Klapperich, Catherine; Cabodi, Mario; Gaydos, Charlotte A.; Rompalo, Anne M.; Manabe, Yukari; Wang, Tza-Huei; Rothman, Richard; Geddes, Chris; Widdice, Lea; Jackman, Joany; Mathura, Rishi A.; Lash, Tiffani Bailey
To advance the development of point-of-care technology (POCT), the National Institute of Biomedical Imaging and Bioengineering established the POCT Research Network (POCTRN), comprised of Centers that emphasize multidisciplinary partnerships and close facilitation to move technologies from an early stage of development into clinical testing and patient use. This paper describes the POCTRN and the three currently funded Centers as examples of academic-based organizations that support collaborations across disciplines, institutions, and geographic regions to successfully drive innovative solutions from concept to patient care.
Metal-enhanced fluorescence from zinc substrates can lead to spectral distortion and a wavelength dependence
(AIP, 2015-02-25) Hamo, Hilla Ben; Karolin, Jan; Mali, Buddha L.; Kushmaro, Ariel; Marks, Robert; Geddes, Chris
Metal-enhanced fluorescence enhancement factors up to 7-fold have been observed for Basic Fuchsin (BF) in close proximity to Zinc nano particulate substrates. In addition, the emission spectra of BF close-to Zinc as compared to a control sample are heavily distorted, particularly on the red-edge, giving systematic trends in enhancement, anywhere from 3- to 7-fold. We discuss these remarkable wavelength dependent effects with regard to the mechanism of metal-enhanced fluorescence.
Microwave-accelerated method for ultra-rapid extraction of Neisseria gonorrhoeae DNA for downstream detection
(Elsevier, 2016-10-01) Melendez, Johan; Santaus, Tonya; Brinsley, Gregory; Kiang, Daniel; Mali, Buddha; Hardick, Justin; Gaydos, Charlotte A.; Geddes, Chris
Nucleic acid-based detection of gonorrhea infections typically require a two-step process involving isolation of the nucleic acid, followed by detection of the genomic target often involving polymerase chain reaction (PCR)-based approaches. In an effort to improve on current detection approaches, we have developed a unique two-step microwave-accelerated approach for rapid extraction and detection of Neisseria gonorrhoeae (gonorrhea, GC) DNA. Our approach is based on the use of highly focused microwave radiation to rapidly lyse bacterial cells, release, and subsequently fragment microbial DNA. The DNA target is then detected by a process known as microwave-accelerated metal-enhanced fluorescence (MAMEF), an ultra-sensitive direct DNA detection analytical technique. In the current study, we show that highly focused microwaves at 2.45 GHz, using 12.3-mm gold film equilateral triangles, are able to rapidly lyse both bacteria cells and fragment DNA in a time- and microwave power-dependent manner. Detection of the extracted DNA can be performed by MAMEF, without the need for DNA amplification, in less than 10 min total time or by other PCR-based approaches. Collectively, the use of a microwave-accelerated method for the release and detection of DNA represents a significant step forward toward the development of a point-of-care (POC) platform for detection of gonorrhea infections.
Phosphorus sequestration in the form of polyphosphate by microbial symbionts in marine sponges
(PNAS, 2015-04-07) Zhang, Fan; Blasiak, Leah C.; Karolin, Jan; Powell, Ryan J.; Geddes, Chris; Hill, Russell T.
Marine sponges are major habitat-forming organisms in coastal benthic communities and have an ancient origin in evolution history. Here, we report significant accumulation of polyphosphate (polyP) granules in three common sponge species of the Caribbean coral reef. The identity of the polyP granules was confirmed by energy-dispersive spectroscopy (EDS) and by the fluorescence properties of the granules. Microscopy images revealed that a large proportion of microbial cells associated with sponge hosts contained intracellular polyP granules. Cyanobacterial symbionts cultured from sponges were shown to accumulate polyP. We also amplified polyphosphate kinase (ppk) genes from sponge DNA and confirmed that the gene was expressed. Based on these findings, we propose here a potentially important phosphorus (P) sequestration pathway through symbiotic microorganisms of marine sponges. Considering the widespread sponge population and abundant microbial cells associated with them, this pathway is likely to have a significant impact on the P cycle in benthic ecosystems.
Extraction and Sensitive Detection of Toxins A and B from the Human Pathogen Clostridium difficile in 40 Seconds Using Microwave-Accelerated Metal-Enhanced Fluorescence
(PLOS, 2014-08-27) Joshi, Lovleen Tina; Mali, Buddha L.; Geddes, Chris; Baillie, Les
Clostridium difficile is the primary cause of antibiotic associated diarrhea in humans and is a significant cause of morbidity and mortality. Thus the rapid and accurate identification of this pathogen in clinical samples, such as feces, is a key step in reducing the devastating impact of this disease. The bacterium produces two toxins, A and B, which are thought to be responsible for the majority of the pathology associated with the disease, although the relative contribution of each is currently a subject of debate. For this reason we have developed a rapid detection assay based on microwave-accelerated metal-enhanced fluorescence which is capable of detecting the presence of 10 bacteria in unprocessed human feces within 40 seconds. These promising results suggest that this prototype biosensor has the potential to be developed into a rapid, point of care, real time diagnostic assay for C. difficile.
In Situ Enzymatic Conversion of Nannochloropsis oceanica IMET1 Biomass into Fatty Acid Methyl Esters
(Springer, 2017-06-01) Wang, Yao; Lee, Yi-Ying; Santaus, Tonya; Newcomb, Charles E.; Liu, Jin; Geddes, Chris; Zhang, Chengwu; Hu, Qiang; Li, Yantao
Conventionally, production of methyl ester fuels from microalgae occurs through an energy-intensive two-step chemical extraction and transesterification process. To improve the energy efficiency, we performed in situ enzymatic conversion of whole algae biomass from an oleaginous heterokont microalga Nannochloropsis oceanica IMET1 with the immobilized lipase from Candida antarctica. The fatty acid methyl ester yield reached 107.7% for dry Nannochloropsis biomass at biomass to t-butanol to methanol weight ratio of 1:2:0.5 and a reaction time of 12 h at 25 °C, representing the first report of efficient whole algae biomass conversion into fatty acid methyl esters at room temperature. Different forms of algal biomass including wet Nannochloropsis biomass were tested. The maximum yield of wet biomass was 81.5%. Enzyme activity remained higher than 95% after 55 days of treatment (equal to 110 cycles of reaction) under the conditions optimized for dry algae biomass conversion. The low reaction temperature, high enzyme stability, and high yield from this study indicate in situ enzymatic conversion of dry algae biomass may potentially be used as an energy-efficient method for algal methyl ester fuel production while allowing co-product recovery.
Metal-enhanced fluorescence from copper substrates
(AIP, 2007-04-25) Zhang, Yongxia; Aslan, Kadir; Previte, Michael J. R.; Geddes, Chris
In this letter, the authors report the observation of metal-enhanced fluorescence emission from fluorophores deposited on copper (Cu) substrates. Different thicknesses of Cu particulate films (from 1 to 5nm) were deposited onto glass slides using thermal vapor deposition. Fluorophores positioned in close proximity to the Cu films show fluorescence enhancement as a function of the Cu thickness increases, reaching a maximum (2.5-fold) at ≈3nm. The findings strongly suggest that surface plasmons from Cu can radiate and therefore enhance a fluorophore’s spectral properties, similar to observations reported for both silver and gold nanoparticle deposited substrates.
Metal Enhanced Fluorescence Solution-based Sensing Platform 2: Fluorescent Core-Shell Ag@SiO₂ Nanoballs
(Springer, 2007-02-06) Aslan, Kadir; Wu, Meng; Lakowicz, Joseph R.; Geddes, Chris
In this Rapid Communication, we present the development of monodisperse core-shell (silver core-silica shell) nanoparticles with various shell thicknesses featuring a fluorophore, subsequently named Metal-Enhanced Fluorescence (MEF) nanoballs. MEF nanoballs consist of a ≈130 nm silver nanoparticle core, a silica shell with up to 35 nm thickness and fluorophores doped within the silica shell. Fluorescent nanobubbles where the silver core is removed by chemical etching are used as control samples to show the benefits of using silver nanoparticles, i.e, Metal-Enhanced Fluorescence. Finally, we demonstrate the broad potential biological applications of MEF nanoballs by employing near-infra red emitting probes (Rhodamine 800) within the silica shell, for potential applications in cellular imaging and solution-based sensing.

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