Fig 1 shows paleochannel locations recognized from planview fluv

Fig. 1 shows paleochannel locations recognized from planview fluvial architectural elements, from visible satellite imagery (LANDSAT, SPOT, DigitalGlobe satellites), and identified from their topographic expression (Syvitski et al., 2012) as reconstructed check details from the SRTM topography (Fig. 2). Channel names (and their spelling) are from Holmes (1968), who applied forensic historical analysis to determine when these channels would have been most active. Holmes (1968) identified three channel patterns expressed within air photos (Fig. 1): circa 325 BC, 900 AD and 1600 AD. These dates represent generalized periods. Historical

maps were analyzed for their spatial geo-location error (Table 1), by digitally identifying towns on geo-referenced maps and comparing them to modern city locations. Maps earlier than 1811 did not have sufficient positioning detail to have their root-mean-square error determined. Few cities lasted across multiple centuries, in part because Indus River avulsions commonly left river settlements without water resources for drinking, agriculture, or transportation. [Note: Sindh towns often changed their spelling AG-014699 chemical structure and towns that were re-located sometimes kept their old name: see supplementary spelling data.] Pinkerton (1811; see suppl. matl.) notes that the Indus River

was navigable from the mouth to the province of Lahore, 900 km upstream for ships of 200 tons. At that time the Indus River system included an extensive set of natural overflow flood pathways across the Indus plain as indicated by Lapie (1829; see suppl. matl.). An SDUK 1838 map shows the Indus flowing on both sides of Bukkur, an island near Sukkur. The same map indicates that the Indus was typically 500 m wide, 12 m deep, with a flow of 1.5 m/s (∼4500 to 9000 m3/s) and rose 4 m during flood (i.e. ∼12,000 to 16,000 m3/s) – values that are similar to those of today. The Western Nara River, a northern offshoot course of the main Indus, originated near Kashmore (Fig. 1) in pre-historic time and later near Ghauspur (Panhwar, 1969). As the Indus moved west, this distributary was

Bumetanide 37 km north of Larkana by 1860 and only 15 km north by 1902, when it was converted into a canal (Panhwar, 1969). Johnston (1861; see suppl. matl.) shows the Eastern Nara River to be a viable secondary pathway of Indus water to the sea through a complex of river channels. In 1859, the Eastern Nara was converted into a perennial canal (Panhwar, 1969). The Indus adopted its present course west of Hyderabad in 1758 when the Nasarpur Course was deserted (Fig. 1) and discharge greatly decreased down the Eastern Nara (Fig. 1) (Wilhelmy, 1967 and Holmes, 1968). The Fuleti River, a significant discharge branch to the west of Hyderabad through the first half of the 19th century (SDUK, 1833 and Johnston, 1861; see suppl. matl.), became a spillway and occupied the channel of the former Ren River (Fig. 1).

g , Loutre and Berger, 2003, de Abreu et al , 2005 and Tzedakis,

g., Loutre and Berger, 2003, de Abreu et al., 2005 and Tzedakis, 2010). However, irrespective of atmospheric CO2 values, this is likely to be an inappropriate analogue because it does selleck products not consider other very significant

anthropogenic forcings on the carbon cycle, nitrogen cycle, atmospheric methane, land use change and alteration of the hydrological cycle, which were not present during MIS 11 but which are very important in the Anthropocene (e.g. Rockström et al., 2009). Studies of Earth’s climate ‘tipping points’ show that nonlinear forcing–response climatic behaviour, leading to state-shifts in many or all of Earth’s systems, can take place under a number of types of forcings, including the biosphere, thermohaline circulation and continental deglaciation (Lenton et al., 2008). It may be that accelerated deglaciation of Greenland

and the west Antarctic see more ice sheet, as result of Anthropocene warming and sea-level rise, will have similar impacts on global thermohaline circulation as deglaciations of the geologic past. However, changes in land surface hydrology and land use may result in a range of unanticipated environmental outcomes that have little or no geologic precedence (e.g. Lenton, 2013). Based on these significant differences between the Anthropocene and the geologic past, we argue that monitoring and modelling climate and environmental change in the Anthropocene requires a new kind of ‘post-normal science’ that cannot lean uncritically on our knowledge of the geological past (e.g., Funtowicz and Ravetz, 1993 and Funtowicz and Ravetz, 1994). In terms of Earth system dynamics, the Anthropocene can be best considered as a singularity in which its constituent Earth systems are increasingly exhibiting uncertainty in the ways in which systems operate. This results in a high degree of uncertainty (low predictability) in the outcome(s)

of forcings caused by direct and indirect human activity. Moreover, climate models and analysis of Earth system dynamics during periods oxyclozanide of very rapid climate and environmental change, such as during the last deglaciation, suggest that very rapid system changes as a result of bifurcations are highly likely (Held and Kleinen, 2004, Lenton, 2011 and Lenton, 2013). This supports the viewpoint that Earth systems in the Anthropocene are likely to be increasingly nonlinear and thus are a poor fit to uniformitarian principles. We argue that understanding and modelling of Earth systems as ‘low-predictability’ systems that exhibit deterministic chaos should be a key goal of future studies.

The tourism infrastructure is dominantly controlled by the Kinh <

The tourism infrastructure is dominantly controlled by the Kinh Veliparib purchase majority, while the other minorities mainly deliver labour force to run the tourism industry. In order to evaluate the potential impact of tourism activities on forest cover in Sa Pa, three land cover maps were compiled based on LANDSAT images available from the U.S. Geological Survey archives (http://glovis.usgs.gov). One LANDSAT-patch (path/row 128/45) covers the whole Sa Pa district with a resolution of 30 m by 30 m. The Landsat images

date from Feb 1, 1993 (just after the opening for international tourism), Nov 4, 2006 (midst of the evaluation period) and Jan 02, 2014 (current state). All images were taken in the post-harvest period when the arable fields are bare. All Landsat images in the freely available USGS archive are orthorectified with precision terrain correction level L1T (Vanonckelen et al., 2013). All images were then corrected for atmospheric and topographic effects using the MODTRAN-4 code and the semi-empirical topographic correction implemented in ATCOR2/3 (Richter, 2011 and Balthazar et al., 2012). Then, a supervised maximum likelihood classification was carried out to map the following 5 land cover categories (Fig. 2): forest, shrub, arable land, water body and urban area. Spectral signatures for the different land cover types were identified

by delineating training areas on the basis of field work C646 carried out in 2010 (Fig. 5). The accuracy of the land cover maps was assessed by comparing the classified land cover with visual interpretations of very high resolution remote sensing data. For 1993, the comparison was done with aerial photographs (MONRE, 1993); for 2006 with a VHR-SPOT4 image (MONRE, 2006) and for 2014 with a VHR-SPOT5 image (MONRE, 2012). Random sampling of validation points was done with n = 219 for the 1993 map, n = 315 for the 2006 map, and n = 306 for the 2014 map. The number of

sample points per land cover class varied from 3 to 111, depending on the areal cover of the classes. For all randomly selected points, the land cover was compared with the classified land cover. This comparison allowed to assess the overall accuracy, quantity disagreement Cyclin-dependent kinase 3 and allocation disagreement (in %) following the procedures described by Pontius and Millones (2011). In order to analyze land cover change trajectories over 3 timeperiods, the change trajectories were grouped in 6 classes: (1) deforestation (change from any class of forest to non-forest), (2) reforestation (change from non-forest to forest), (3) land abandonment (change from agricultural land to shrub or forest), (4) expansion of arable land (conversion from shrub to arable land), (5) other changes, and (6) no change (Table 1). The original classes ‘water body’ and ‘urban area’ that only occupy a minor fraction of the land were not taken into consideration.

1% of the patients were ≤49 years, and 41 2% were ≥60 years Unfo

1% of the patients were ≤49 years, and 41.2% were ≥60 years. Unfortunately, we did not obtain any conclusive labeling for ZD1839 order MGMT (instead, the controls were positive), though we used a robust antibody (SPM287). In fact, the small tissue cores (1.0 mm) and the well-known MGMT immunolabeling heterogeneity may have been limiting factors in our analysis, underscoring some of the difficulties in using immunohistochemistry to assess MGMT expression in formalin-fixed paraffin-embedded GBM tissues, as previously reported

in other studies [34] and [46]. Similarly, the immunohistochemistry for IDH1 was negative in all GBM tissue cores (with positive controls). However, it is important to note that the majority (if not all) of our GBM cases were primary GBMs that did not contain the IDH1 mutation. Although we used a general IDH1-antibody instead of the well-established antibody for the dominant mutant variant of the enzyme (IDH1-R132H), find more we do not believe that it impacted our results because no IDH1-immunopositive cells could be found in the TMAs. Furthermore, the staining of such small areas with the mutation-specific antibody may be problematic. In conclusion, 50.5% of the glioblastomas expressed variable levels of FasL, 68.9% expressed Fas, 45.7% expressed cleaved caspase-8,

and 35.2% expressed cleaved caspase-3. Moreover, glioblastoma tumors should contain a functional mechanism for the extrinsic apoptotic pathway. Our findings suggest that Fas–Fas-ligand downstream signal transduction could be inhibited, especially at the stage of caspase-8 activation, thereby establishing a major mechanism for the evasion of apoptosis by these tumors. Furthermore, our findings highlight the study of Ho et al. [16], who showed that FasL and Fas delivery by a glioma-specific and cell cycle-dependent HSV-1 amplicon virus enhances

apoptosis in high-grade gliomas, and may be useful as an adjuvant therapy to complement the current therapeutic regimens for human gliomas. In addition, the low immunoexpression of cleaved caspase-8 (0 to <50% of faintly positive tumor cells) in glioblastomas was an independent about prognosticator of slightly decreased disease-specific survival, compared with tumors that expressed higher levels of cleaved caspase-8. Further studies examining molecular targets in the extrinsic pathway of apoptosis are needed and may reveal promising treatment strategies for glioblastomas. The authors declare that there are no conflicts of interest. We would like to thank Joaquim Soares de Almeida, who prepared the tissue microarrays, and Maria José Carregosa Pinheiro dos Santos for their excellent technical assistance. This work was supported by Fundação de Amparo a Pesquisa do Estado de São Paulo-FAPESP (04/09932-4). Writing assistance was provided by BioMed Proofreading, Cleveland, USA.

In this case, the wall was deformed A large superficial flat neo

In this case, the wall was deformed. A large superficial flat neoplasm was the cause

of this deformity. Figure options Download full-size image Download high-quality image (609 K) Download as PowerPoint slide Fig. 16. General to detailed visualization of a superficial elevated neoplasm and its imaging documentation. Examination of a lesion to understand the significance of its detail is a fluid stepwise process. For example, (A) on detection, the lesion is first viewed in a long view, to understand and evaluate its relative size, shape, and location. The lesion is then examined with varying expansion of the colon. Increasing (B) or decreasing (C) air insufflation may help improve visualization of a flat or depressed lesion. (D) Closer view permits detailed examination of the vessel and surface pattern. (E, F) Application of indigo carmine BI 2536 chemical structure dye further enhances the borders of the Trichostatin A lesion and the details of the morphology and surface pattern. Figure options Download full-size image Download high-quality image (318

K) Download as PowerPoint slide Fig. 17. General to detailed visualization of a flat neoplasm and its imaging documentation, illustrating the use of a translucent distal attachment device (cap) in the detailed view and understanding of the lesion. Documentation of the lesion is best performed by taking an overview (long-shot) picture, before close-up pictures are taken (A, B, C). In (A), the lesion is inspected using high definition white light. In (B), narrow-band imaging (NBI) was used to visualize the surface and microvessel patterns. In (C), indigo carmine was used to determine the margin of the lesion. Pit-pattern

characterization of the lesion using either NBI or indigo carmine is generally not useful. Detailed imaging of the lesion is critical for its complete resection. (D) A circumferential cut was performed to isolate the lesion before its snaring. Figure options Download full-size image Download high-quality image (238 K) Download as PowerPoint slide Fig. 18. (A–C) White-out (halation) can impair adequate viewing and interpretation. There is a blurred effect around the edges of the area highlighted caused by reflection and scattering of light. Figure options Download full-size image Download high-quality image (343 K) Uroporphyrinogen III synthase Download as PowerPoint slide Fig. 19. Appropriate setting of the iris is important. The iris function on endoscope processors adjusts the distribution of light, and is generally sufficient to adjust brightness. • Auto: The brightness is adjusted based on the brightest part of the central part and the average brightness of the periphery part. Figure options Download full-size image Download high-quality image (301 K) Download as PowerPoint slide Fig. 20. Inadequate documentation and preparation, and inappropriate use of, image-enhanced endoscopy. A picture is worth a thousand words, except when the picture is not adequate.

However, as one of the predicted carboxypeptidases A (contig 48)

However, as one of the predicted carboxypeptidases A (contig 48) has a predicted GPI-anchor, it is highly probable that the membrane-bound activity is a truly microvillar protein, whereas the soluble ones are released by microapocrine secretion. Six lipases are similar to pancreatic lipases and five are supposed

to be released by microapocrine secretion. One of the pancreatic lipases (contig 379) has a puzzling predicted transmembrane loop. Only one gastric lipase (contig 673) was found in microapocrine vesicles. Except for proteins thought to be part of the secretory machinery and transporters, other predicted proteins that are secreted by microapocrine secretion MLN0128 ic50 are listed in Table 4, in spite of lacking data on signal peptides. Most putative secretory proteins (aminopeptidase, Metformin clinical trial carboxyl esterase, prolyl carboxypeptidase, lipase, and

serine protease) are digestive enzymes with few proteins involved in protection and PM. The ATPases (contigs 435 and 500) are probably coding for proton pumps that acidify the vesicle contents as is usual in secretory vesicles (Alberts et al., 2008). The organic cation transporter (contig 631) may derive from the microvillar membrane, although there is no experimental support for this claim. Predicted proteins that are supposed to be involved in the secretory machinery are listed in Table 2 and Table 4. The predicted proteins calmodulin, annexin, myosin 7a and, gelsolin 1 are not anchored. They might be recovered in the microvillar membrane preparations

because putatively they associate with membranes or with cystoskeleton elements found contaminating the preparations. Calmodulin, annexin, myosin Selleckchem 5-FU 7a, and gelsolin 1 putatively interplay in the microapocrine secretory process of digestive enzymes described in S. frugiperda midgut ( Ferreira et al., 1994, Jordão et al., 1999, Bolognesi et al., 2001 and Ferreira et al., 2007) but further work is necessary to settle this subject. This work was supported by the Brazilian research agencies FAPESP (Temático) and CNPq. We are indebted to W. Caldeira, and M.V. Cruz for technical assistance. W. Silva is a doctoral fellow of CAPES. C. Ferreira and W.R. Terra are staff members of their respective department, research fellows of CNPq, and members of the Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular. “
“The juvenile period is a time of intensive nutrient uptake that supports insect growth and transition to adult morphology and metabolism. Food ingestion is specially intense among Lepidoptera as their feeding is mainly restricted to plants, which are a poor sources of nutrients (Dow et al., 1987 and Klowden, 2007). During digestion, nutrients are mobilized by a set of hydrolases (Terra and Ferreira, 2005 and Terra and Ferreira, 1994) and posteriorly absorbed by several transporters (Meleshkevitch et al., 2006 and Meleshkevitch et al., 2009) using the so-called “voltage strategy” (Harvey and Okech, 2010).

These particles ERK

These particles selleck kinase inhibitor have the additional advantage that they also allow the determination of the deposition rate in their core-fluorescently labeled form. Multi-walled carbon nanotubes with different diameters were used to identify the influence of shape and the aggregation behavior of the nanoparticles since carbon nanotubes show high aggregation and polystyrene particles low aggregation (Wiesner and Colvin, 2005). Carbon nanotubes are also candidates for numerous medical applications (Zhang et al., 2010). 20, 40, 100, and 200 nm red (580/605) fluorescent labeled carboxyl-functionalized polystyrene particles (FluoSpheres)

were purchased from Invitrogen (Vienna). Carboxylated short multi-walled carbon nanotubes (0.5–2 μm long, purity >95%) with outer diameters <8, 20–30 and >50 nm were obtained from CheapTubes Inc. (Brattleboro, Vermont). To identify a potential difference in cytotoxicity between exposure in submersed culture and exposure as aerosol, 20 nm amine-functionalized polystyrene particles were used (Estapor Microspheres, Merck Chimie S.A.S., Fontenay-sous-Bois). For exposure all nanoparticles were diluted and the suspensions were put into

an Elmasonic Dabrafenib S40 water bath (ultrasonic frequency: 37 kHz, Elma, Singen) for 20 min prior to all experiments. For the VITROCELL system and the MicroSprayer cells were cultured for 24 h after the exposures. Fluorescein sodium salt (Sigma Aldrich, Steinheim) was used as a macromolecular reference substance. Nanoparticle-sizes were determined routinely C1GALT1 by dynamic light scattering using a Zetasizer Nano ZS (Malvern Instruments, Malvern) equipped

with a 532 nm HeNe laser, taking into account viscosity as well as refraction index. Polystyrene particles were diluted with the same solvent used for the exposures (distilled water for VITROCELL PT/PARI BOY LC Sprint system and DMEM + 10% FBS for MicroSprayer) to a concentration of 200 μg/ml and sonicated for 20 min before measurements. To study the stability of the nanoparticles in the VITROCELL PT/PARI BOY LC Sprint system samples of the condensate from the vial at the end of the glass tube (Fig. 1a) were also tested. After equilibration of the sample solution to 25 °C, scattered light was detected at a 173° angle with laser attenuation and the dynamic fluctuations of light scattering intensity caused by Brownian motion of the particles was evaluated. Polydispersity Indices <0.2 are interpreted as indication for homogenous samples (Stancampiano et al., 2008). The viscosities of the different solutions were 0.88 cP (water) and 0.94 cP (DMEM + 10% FBS). Additionally, the refraction indices of all investigated media were found to be around 1.33 (1.33 for water, 1.345 DMEM + 10% FBS. These values were very similar and showed no impact on the measured particle sizes. CNTs were suspended in DMEM + 10% FBS at 1 mg/ml.

Management capacity varied greatly among the

13 fishery a

Management capacity varied greatly among the

13 fishery agencies, especially in the number of export inspection officers, number of scientists with skills in stock assessment and patrol boats for inspections at sea (Fig. 2). Micronesian countries have weaker capacity for managing sea cucumber fisheries than most agencies in Melanesia and Polynesia. Concerning the Micronesian countries, none had skilled officers LY2109761 purchase to conduct stock assessment analyses, they had fewer officers who could identify sea cucumber species than in Melanesian and Polynesian countries, none had funding for underwater visual censuses, and none had patrol boats for inspectiing sea cucumbers at sea. Technical capacity in fishery agencies was relatively strong for some management tasks and weak for others. The number of agency scientists with technical skills in stock assessment (e.g. to calculate maximum sustainable yield) varied widely among the 13 fisheries. Half of the countries had no such

scientists. Management agencies generally had many officers (average=6) responsible for planning and implementing marine reserves. All but two agencies had at least three officers who can identify live sea cucumbers to species level. On the other hand, just 5 of the 13 agencies had more than two officers selleckchem trained in export inspections and one quarter of countries have no trained inspection officers. More than three quarters (79%) of fishery agencies have human resources and skills for underwater visual census (UVC) but, paradoxically, less than one quarter (21%) has funding for conducting regular UVCs. All but three fishery managers reported difficulty in obtaining monthly information on catch from fishers. Enforcement and inspection Thiamet G capacity was generally very weak. On average, agencies have less than two boats for inspections at sea and half of them have none. Half of the managers believed that landings of (fresh) sea cucumbers are

checked “practically never” in their fishery. Sea cucumber landings were checked one or more times per week in only four fisheries. In most cases, bags of beche-de-mer (dried sea cucumbers) are checked occasionally prior to export, and in four of the export fisheries they are checked “regularly”. In just half of the export fisheries, inspection officers have received training in identifying dried sea cucumbers. More than two out of three (71%) government agencies had not established formal management objectives for their sea cucumber fisheries and most (79%) did not have reference points for assessing management performance. During the workshop, the 10 multi-disciplinary management objectives were ranked quite differently among the fishery managers (Fig. 3). The objective ranked most important, on average, was to maintain stocks at levels to sustain viable populations and recruitment.

Male Swiss mice weighing 18–22 g were used The animals were main

Male Swiss mice weighing 18–22 g were used. The animals were maintained for 2 days at the laboratory before experiments with water and food ad libitum in appropriate environmental conditions and were used under ethical conditions. All experimental procedures followed the ethical parameters proposed by the International Society of Toxinology and the Brazilian College of Experimental Animals and were approved by Ethical Committee for Use of Animals of Butantan Institute (protocol n° 591/09). A pool of lyophilized venom,

obtained from various adult specimens of CH5424802 cell line C. durissus terrificus snakes, was supplied by the Laboratory of Herpetology, Butantan Institute. The venom was stored at −20 °C and solutions (w/v) were prepared in sterile saline immediately before use. The crude venom was fractionated in a Mono-Q HR 5/5 column in a FPLC system (Pharmacia, Uppsala, Sweden) as previously described by Rangel-Santos et al. (2004). Three fractions (frI, frII and frIII) were obtained, and frII corresponded to pure crotoxin. The homogeneity of this toxin was checked by non-reducing sodium dodecyl sulfate–polyacrylamide gel electrophoresis (12.5%) (Laemmli, 1970). Crotoxin was also tested for lethality and phospholipase A2 activity (Santoro et al., 1999). The Cdt fractions used throughout this see more study were generously supplied by Dr. Maisa Spendore Della Casa (Laboratory of Immunopathology, Butantan Institute). The BCG used as a phlogistic agent was prepared

with live attenuated bacilli of Mycobacterium bovis (Moreau strain), supplied in lyophilized form by Instituto Butantan. A suspension containing 8 × 105 bacilli in 30 μL of saline solution were injected into the footpad of mice. The contralateral paw received the same volume of saline solution. The concentration of BCG used in this study was based on data from the literature ( Moura and Mariano, 1996). Paw edema was evaluated once a day with the aid of a micrometer (Mitutoyo, Japan), for the 15 days after the BCG injection. In some experiments, edema was evaluated 1, 2, 4 and 6 h after the BCG injection. Results were calculated

as the difference Molecular motor in thickness of both BCG- and saline-injected paws, and edema was expressed as the percentage increase in paw thickness. To identify the inhibitory effect of the C. durissus terrificus venom on chronic paw edema induced by BCG, mice were injected with a single dose (75 μg/kg) of Cdt crude venom subcutaneously (s.c.) in the back 1 h before receiving BCG into the footpad. The paw edema was compared to that obtained in control animals injected with the saline solution (100 μL) instead of the Cdt venom, by the same route (s.c.). To determine if Cdt venom has an inhibitory effect after the initiation of a chronic inflammatory process, mice received an injection of BCG in the footpad. Groups then received Cdt venom in the back (s.c.) 1 h, 6 days or 11 days after the inoculation of BCG. The respective control groups received saline instead of venom.

Purified κ and λ FLC calibrator materials were separately incubat

Purified κ and λ FLC calibrator materials were separately incubated with biotinamidohexanoyl-6-aminohexanoic acid, N-hydroxysuccinimide ester in dimethyl sulfoxide overnight (all Sigma

Aldrich). Biotinylated light chains were then separated using NAP™ 5 Columns (Sephadex G-25 DNA grade; GE Healthcare), eluted in PBS, and the concentration of the eluate was measured by spectrophotometry. After the addition of 0.099% sodium azide, biotinylated light chains were stored at 4 °C, until required. Prior to assaying, each of the anti-κ FLC and anti-λ FLC mAbs was covalently coupled to four different 5.5 μm polystyrene Xmap® beads (Bio-Rad UK) using a two step carbodiimide reaction protocol. Specifically, the different bead regions used were #27 (BUCIS 01), Trichostatin A purchase #28 (BUCIS 04), #29 (BUCIS 03), and #30 (BUCIS 09). After sonication and vortexing, beads were incubated with N-hydroxysuccinimide (NHS) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) for 30 min in activation buffer (0.1 M PBS, pH 6.2). Following two

wash procedures, the bead pellet was incubated for 3 h with the relevant mAbs (100 μL at 1 mg/mL) on a rotor, in the dark, at room temperature. Beads were then washed twice, and finally, the pellet was resuspended in blocking/storage buffer (0.1 M PBS, 0.05% Tween20, 1% BSA, 0.05% NaN3, pH 7.2). The beads selleck chemicals llc were enumerated using a haemocytometer to ensure consistency between conjugations. Labelled beads were stored in blocking/storage buffer in the dark at 4 °C until required Tideglusib in the assay; the beads were found to be stable for at least 6 months in these conditions (data not shown). The efficiency of mAb conjugation to each beadset was determined in a one-step assay by incubating each beadset with goat anti-mouse IgG labelled with phycoerythrin (PE; Southern Biotech, USA), and subsequent measurement by Luminex®. The effectiveness of each mAb-bead complex at detecting κ and λ FLCs was tested in a two-step assay by (1) incubating beads with biotinylated purified κ and λ FLCs, and (2) incubation with streptavidin-PE (Invitrogen, UK), and subsequent

measurement by Luminex®. A minimum median fluorescent intensity (mfi) was set at > 10,000 mfi units for mAb conjugation to beads and > 5000 mfi units for detection of κ and λ FLCs, as this provided calibration curves with the most sustained linearity, and, hence, more reliable and reproducible results on patient samples. Human κ and λ FLCs were measured in a multi-plex competitive inhibition format. 40 μl of biotinylated FLC diluted 1 in 400 in FLC buffer (PBS, 12.5% 2 M Tris, 1% BSA, 0.099% NaN3, 0.05% Tween20) was added to each well in a 1.2 μm MV Multiscreen (Millipore, UK) 96-well filter plate, followed by 40 μl of each sample, calibrators or controls, and then incubated for 30 min with mAb coupled beads. Each sample was diluted 1 in 5 in assay buffer (PBS, 1% BSA, 0.