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Biomedical subjects

E Calvo

Publications and source records attributed to E Calvo.

At least 19 recordsLinked to original sources

Osteoporosis increases the severity of cartilage damage in an experimental model of osteoarthritis in rabbits.

OBJECTIVE: To evaluate the effect of osteoporosis (OP) in cartilage damage developed in an experimental model of osteoarthritis (OA) in mature female rabbits in order to investigate the relationship between OP and OA. METHODS: OA was experimentally induced by anterior cruciate section and partial medial meniscectomy in the left knee of 12 rabbits. OP was experimentally induced prior to OA in six rabbits by bilateral ovariectomy (OVX) and systemic corticosteroid administration during 4 weeks. Knees were evaluated with high resolution magnetic resonance imaging (MRI) before knee surgery to rule out any detrimental effect of corticosteroids on cartilage. Gross and microscopic cartilage changes were assessed 16 weeks after surgery in bilateral knees. Left knees were considered osteoarthritic or osteoarthritic plus osteoporotic. Right knees were used as osteoporotic and healthy controls, respectively. Bone mineral density (BMD) was measured with dual energy X-ray absorptiometry (DXA) at the lumbar spine, global knee and subchondral knee bone, and its variations correlated with cartilage abnormalities. RESULTS: MRI before knee surgery disclosed no cartilage or bone abnormalities in any of the studied groups. OP increased the severity of cartilage abnormalities in experimental knee OA significantly (P<0.05). Cartilage damage was inversely correlated with BMD variations measured at the lumbar spine (r=-0.74; P=0.015). BMD changes in global and subchondral knee bone also showed a trend to correlate inversely with cartilage damage. CONCLUSIONS: Prior induction of OP increases the severity of cartilage damage in experimental OA. Increase in cartilage damage correlates with bone loss. These findings suggest a direct relationship between OP and OA.

Absorptiometry, Photon↗

Expression of the peptide C4b-binding protein beta in the arthritic joint.

BACKGROUND: C4b-binding protein (C4BP) is a plasma oligomeric glycoprotein that participates in the regulation of complement and haemostasis. Complement-regulatory activity depends on the C4BPalpha-polypeptide, whereas the C4BPbeta-polypeptide inactivates protein S, interfering with the anti-coagulatory protein C-dependent pathway. OBJECTIVE: To investigate the expression of C4BPbeta in the rheumatoid joint. METHODS: Expression of C4BP was studied in synovial explants from patients with rheumatoid arthritis, osteoarthritis and healthy controls, using immunohistochemistry and in situ hybridisation. C4BP isoforms and free C4BPbeta were studied in synovial effusions from patients with rheumatoid arthritis, osteoarthritis and microcrystalline arthritis (MCA) by immunoblotting; total and free protein S levels were studied by enzyme immunoassay. RESULTS: C4BPbeta was overexpressed in the synovial membranes of patients with rheumatoid arthritis, in close association with the severity of synovitis and the extension of interstitial fibrin deposits. As many as 85% fluids from patients with rheumatoid arthritis contained free C4BPbeta, whereas this unusual polypeptide was present in 50% fluids from patients with MCA and 40% fluids from patients with osteoarthritis. Free protein S at the effusions was pathologically reduced in patients with rheumatoid arthritis and MCA, and remained normal in patients with osteoarthritis. CONCLUSION: C4BPbeta is expressed by the inflamed synovial tissue, where it can participate in processes of tissue remodelling associated with invasive growth.

Adult↗

Long term NSAID treatment inhibits COX-2 synthesis in the knee synovial membrane of patients with osteoarthritis: differential proinflammatory cytokine profile between celecoxib and aceclofenac.

OBJECTIVE: To compare the effect of celecoxib with that of a classic non-steroidal anti-inflammatory drug (NSAID) on synovial inflammation and on the synovial expression of proinflammatory genes in patients with knee osteoarthritis (OA). METHODS: 30 patients with severe knee OA scheduled for total knee replacement surgery were included in a 3 month clinical trial. They were randomised to two groups: patients treated with celecoxib (CBX) (200 mg/24 h) and patients treated with aceclofenac (ACF) (100 mg/12 h). Those patients with OA who did not want to be treated with NSAIDs served as a control group. During knee surgery, synovial fluid (SF) and synovial membrane (SM) were collected. A SM specimen was fixed and embedded in paraffin and another part was frozen for molecular biology studies. RESULTS: At the end of study both CBX and ACF treated patients showed a significant improvement in pain and knee function compared with controls. Both drugs significantly reduced prostaglandin E(2) (PGE(2)) SF concentration and down regulated COX-2 mRNA and protein expression at the SM. However, synovial macrophage infiltration (CD68 antigen staining) and expression of proinflammatory mediators, such as interleukin 1beta and tumour necrosis factor alpha, were decreased only by CBX treatment. CONCLUSION: Both drugs improved joint pain and function, inhibited SF PGE(2) concentration, and induced a decrease in synovial COX-2 expression and synthesis not related to the tissue inflammatory status. These data suggest that PGE(2) blocking agents may decrease PGE(2) production not only by direct COX-2 inhibition but also by down regulating COX-2 expression and synthesis. However, CBX and ACF appear to have different anti-inflammatory profiles in controlling OA synovial macrophage infiltration and proinflammatory expression.

Aged↗

Molecular biology of testicular germ cell tumors.

Testicular germ cell tumors (TCGT) comprise a heterogeneous group of neoplasms, although all of them are originated from common precursors related to germ cell lineage. Understanding of normal development of germinal cells is essential to define new markers for diagnosis, prognostic subgroups and targeted therapies. Recent advances related to cytogenetic and molecular features have established the role of immunohistochemistry of c-kit, OCT-3/4 and determination of gain of chromosome 12 in the daily workup of premalignant lesions and invasive tumors. This review summarizes the current knowledge in the field of molecular biology of TGCT.

Biomarkers, Tumor↗

Genome-wide analysis of gene expression in adult Anopheles gambiae.

With their genome sequenced, Anopheles gambiae mosquitoes now serve as a powerful tool for basic research in comparative, evolutionary and developmental biology. The knowledge generated by these studies is expected to reveal molecular targets for novel vector control and pathogen transmission blocking strategies. Comparisons of gene-expression profiles between adult male and nonblood-fed female Anopheles gambiae mosquitoes revealed that roughly 22% of the genes showed sex-dependent regulation. Blood-fed females switch the majority of their metabolism to blood digestion and egg formation within 3 h after the meal is ingested, in detriment to other activities such as flight and response to environment stimuli. Changes in gene expression are most evident during the first, second and third days after a blood meal, when as many as 50% of all genes showed significant variation in transcript accumulation. After laying the first cluster of eggs (between 72 and 96 h after the blood meal), mosquitoes return to a nongonotrophic stage, similar but not identical to that of 3-day-old nonblood-fed females. Ageing and/or the nutritional state of mosquitoes at 15 days after a blood meal is reflected by the down-regulation of approximately 5% of all genes. A full description of the large number of genes regulated at each analysed time point and each biochemical pathway or biological processes in which they are involved is not possible within the scope of this contribution. Therefore, we present descriptions of groups of genes displaying major differences in transcript accumulation during the adult mosquito life. However, a publicly available searchable database (http://www.angagepuci.bio.uci.edu/) has been made available so that detailed analyses of specific groups of genes based on their descriptions, functions or levels of gene expression variation can be performed by interested investigators according to their needs.

Aging↗

Bone mineral measurements of subchondral and trabecular bone in healthy and osteoporotic rabbits.

INTRODUCTION: Experimental models of osteoporosis in rabbits are useful to investigate anabolic agents because this animal has a fast bone turnover with predominant remodelling over the modelling processes. For that purpose, it is necessary to characterize the densitometric values of each type of bony tissue. OBJECTIVE: To determine areal bone mass measurement in the spine and in trabecular, cortical and subchondral bone of the knee in healthy and osteoporotic rabbits. DESIGN: Bone mineral content and bone mineral density were measured in lumbar spine, global knee, and subchondral and cortical bone of the knee with dual energy X-ray absorptiometry using a Hologic QDR-1000/W densitometer in 29 skeletally mature female healthy New Zealand rabbits. Ten rabbits underwent triplicate scans for evaluation of the effect of repositioning. Osteoporosis was experimentally induced in 15 rabbits by bilateral ovariectomy and postoperative corticosteroid treatment for 4 weeks. Identical dual energy X-ray absorptiometry (DXA) studies were performed thereafter. RESULTS: Mean values of bone mineral content at the lumbar spine, global knee, subchondral bone and cortical tibial metaphysis were: 1934+/-217 mg, 878+/-83 mg, 149+/-14 mg and 29+/-7.0 mg, respectively. The mean values of bone mineral density at the same regions were: 298+/-24 mg/cm(2), 455+/-32 mg/cm(2), 617+/-60 mg/cm(2) and 678+/-163 mg/cm(2), respectively. Bone mineral content and bone density of healthy rabbits followed a normal distribution at the four skeletal regions studied. Precision after triplicate repositioning yielded a coefficient of variation ranging from 2.6% to 3.8%. The least significant change ranged between 7.3% and 10.7%. Bone mineral density measured at the four different skeletal regions correlated significantly. Bone mineral density in osteoporotic rabbits was significantly lower in the four regions studied than that in controls, rendering a T-score of, respectively, -2.0+/-1.1 in the lumbar spine, -2.2+/-2.1 in the global knee, -1.9+/-0.6 in the subchondral bone, and -5.7+/-3.1 in the cortical tibia (P<0.05). CONCLUSIONS: DXA is a reliable and precise method to evaluate the bone mass in rabbits. Our results also suggest that subchondral bone is a bone of mixed densitometric characteristics with marked cortical bone predominance.

Absorptiometry, Photon↗

SERS in PAH-Os and gold nanoparticle self-assembled multilayers.

We present a detailed structural and surface-enhanced Raman scattering (SERS) study of poly(allylamine) modified with Os(byp)2ClPyCHO (PAH-Os) and gold nanoparticles self-assembled multilayers [PAH-Os+(Au-nanoparticlesPAH-Os)n, n=1 and 5]. Atomic force microscopy and variable-angle spectroscopic ellipsometry measurements indicate that the first nanoparticle layer grows homogenously by partially covering the substrate without clustering. Analyzing the sample thickness and roughness we infer that the growth process advances thereafter by filling with nanoparticles the interstitial spaces between the previously adsorbed nanoparticles. After five immersion steps the multilayers reach a more compact structure. The interaction between plasmons of near-gold nanoparticles provides a new optical absorption around 650 nm which, in addition, allows a more effective SERS process in that spectral region than at the single-plasmon resonance (approximately 530 nm). We compare the electronic resonance Raman and SERS amplification mechanisms in these self-assembled multilayers analyzing Raman resonance scans and Raman intensity micromaps. As a function of nanoparticle coverage we observe large changes in the Raman intensity scans, with maxima that shift from the electronic transitions, to the plasmon resonance, and finally to the coupled-plasmon absorption. The Raman micromaps, on the other hand, evidence huge intensity inhomogeneities which we relate to "hot spots." Numerical discrete dipole approximation calculations including the interaction between gold nanoparticles are presented, providing a qualitative model for the coupled-plasmon absorption and redshifted Raman hot spots in these samples.

Journal Article↗

Randomized phase II trial of carboplatin versus paclitaxel and carboplatin in platinum-sensitive recurrent advanced ovarian carcinoma: a GEICO (Grupo Espanol de Investigacion en Cancer de Ovario) study.

BACKGROUND: The aim of this study was to determine whether the response rate for the paclitaxel-carboplatin combination is superior to carboplatin alone in the treatment of patients with platinum-sensitive recurrent ovarian carcinoma. PATIENTS AND METHODS: Patients with recurrent ovarian carcinoma, 6 months after treatment with a platinum-based regimen and with no more than two previous chemotherapy lines, were randomized to receive carboplatin area under the curve (AUC) 5 (arm A) or paclitaxel 175 mg/m(2) + carboplatin AUC 5 (arm B). The primary end point was objective response, following a 'pick up the winner' design. Secondary end points included time to progression (TTP), overall survival, tolerability and quality of life (QoL). RESULTS: Eighty-one patients were randomized and included in the intention-to-treat analysis. The response rate in arm B was 75.6% [26.8% complete response (CR) + 48.8% partial response (PR)] [95% confidence interval (CI) 59.7% to 87.6%] and 50% in arm A (20% CR + 30% PR) (95% CI 33.8% to 66.2%). No significant differences were observed in grade 3-4 hematological toxicity. Conversely, mucositis, myalgia/arthralgia and peripheral neurophaty were more frequent in arm B. Median TTP was 49.1 weeks in arm B (95% CI 36.9-61.3) and 33.7 weeks in arm A (95% CI 25.8-41.5). No significant differences were found in the QoL analysis. CONCLUSIONS: Paclitaxel-carboplatin combination is a tolerable regimen with a higher response rate than carboplatin monotherapy in platinum-sensitive recurrent ovarian carcinoma.

Adult↗

Reproducibility, bioinformatic analysis and power of the SAGE method to evaluate changes in transcriptome.

The serial analysis of gene expression (SAGE) method is used to study global gene expression in cells or tissues in various experimental conditions. However, its reproducibility has not yet been definitively assessed. In this study, we have evaluated the reproducibility of the SAGE method and identified the factors that affect it. The determination coefficient (R2 ) for the reproducibility of SAGE is 0.96. However, there are some factors that can affect the reproducibility of SAGE, such as the replication of concatemers and ditags, the number of sequenced tags and double PCR amplification of ditags. Thus, corrections for these factors must be made to ensure the reproducibility and accuracy of SAGE results. A bioinformatic analysis of SAGE data is also presented in order to eliminate these artifacts. Finally, the current study shows that increasing the number of sequenced tags improves the power of the method to detect transcripts and their regulation by experimental conditions.

Animals↗

Transient bone marrow oedema of the knee.

OBJECTIVE: To evaluate the role of MRI in the diagnosis of transient bone marrow oedema of the knee. DESIGN: All cases were analysed with plain film and MR. Bone scan was performed to one of them. Laboratory tests were obtained in all 8 cases and core decompression was performed in two cases. PATIENTS: The study includes 8 patients, 7 men and 1 woman with ages ranging from 33 to 74 years with severe knee pain, tenderness and slightly limited range of motion. RESULTS AND CONCLUSIONS: Radiographs were unremarkable in terms of osteopenia because they were obtained early in the course of the disease and proved to be of no value in the diagnosis. MR is the imaging modality of choice showing low signal intensity in T1WI and increased signal intensity in T2WI with no joint destruction, fractures, bone death, periostitis or cortical disruption. Bone scan showed increase uptake of the affected area. Laboratory tests were normal. Spontaneous resolution was the rule. It should not be a diagnosis of exclusion and it should be ruled out every time a patient presents with pain and limited range of motion in the absence of history of trauma.

Adult↗

Phosphorylation and subcellular localization of transmissible gastroenteritis virus nucleocapsid protein in infected cells.

The nucleocapsid (N) protein is the only phosphorylated structural protein of the coronavirus Transmissible gastroenteritis virus (TGEV). The phosphorylation state and intracellular distribution of TGEV N protein in infected cells were characterized by a combination of techniques including: (i) subcellular fractionation and analysis of tryptic peptides by two-dimensional nano-liquid chromatography, coupled to ion-trap mass spectrometry; (ii) tandem mass-spectrometry analysis of N protein resolved by SDS-PAGE; (iii) Western blotting using two specific antisera for phosphoserine-containing motifs; and (iv) confocal microscopy. A total of four N protein-derived phosphopeptides were detected in mitochondria-Golgi-endoplasmic reticulum-Golgi intermediate compartment (ERGIC)-enriched fractions, including N-protein phosphoserines 9, 156, 254 and 256. Confocal microscopy showed that the N protein found in mitochondria-Golgi-ERGIC fractions localized within the Golgi-ERGIC compartments and not with mitochondria. Phosphorylated N protein was also present in purified virions, containing at least phosphoserines 156 and 256. Coronavirus N proteins showed a conserved pattern of secondary structural elements, including six beta-strands and four alpha-helices. Whilst serine 9 was present in a non-conserved domain, serines 156, 254 and 256 were localized close to highly conserved secondary structural elements within the central domain of coronavirus N proteins. Serine 156 was highly conserved, whereas no clear homologous sites were found for serines 254 and 256 for other coronavirus N proteins.

Amino Acid Sequence↗

Microarray analysis of genes showing variable expression following a blood meal in Anopheles gambiae.

A microarray analysis of 14 900 genes of the malaria vector mosquito, Anopheles gambiae, shows that as many as 33% (4924) of their corresponding transcription products vary in abundance within 24 h after a blood meal. Approximately half (2388) of these products increase in their accumulation and the remainder (2536) decrease. Expression dynamics of 80% of the genes analysed by expressed sequence tag (EST) projects reported previously are consistent with the observations from this microarray analysis. Furthermore, the microarray analysis is more sensitive in detecting variation in abundance of gene products expressed at low levels and is more sensitive overall in that a greater number of regulated genes are detected. Major changes in transcript abundance were seen in genes encoding proteins involved in digestion, oogenesis and locomotion. The microarray data and an electronic hyperlinked version of all tables are available to the research community at http://www.angagepuci.bio.uci.edu/1/.

Animals↗

Criteria for arthroscopic treatment of anterior instability of the shoulder: a prospective study.

We prospectively evaluated 61 patients treated arthroscopically for anterior instability of the shoulder at a mean follow-up of 44.5 months (24 to 100) using the Rowe scale. Those with post-operative dislocation or subluxation were considered to be failures. Logistic regression analysis was used to identify patients at increased risk of recurrence in order to develop a suitable selection system. The mean Rowe score improved from 45 pre-operatively to 86 at follow-up (p < 0.001). At least one episode of post-operative instability occurred in 11 patients (18%), although their stability improved (p = 0.018), and only three required revision. Subjectively, eight patients were dissatisfied. Age younger than 28 years, ligamentous laxity, the presence of a fracture of the glenoid rim involving more than 15% of the articular surface, and post-operative participation in contact or overhead sports were associated with a higher risk of recurrence, and scored 1, 1, 5 and 1 point, respectively. Those patients with a total score of two or more points had a relative risk of recurrence of 43% and should be treated by open surgery.

Adolescent↗

Resonant Raman spectroscopy of PAH--Os self-assembled multilayers.

We present a resonant Raman scattering study of (PAH--Os/PVS)n and (PAH--Os/GOx)m self-assembled multilayers (n=1-11 and m=1-3). These Os polymer multilayers can be used in electrodes as efficient molecular wires for biomolecular recognition. The Raman intensity dependence on the number of self-assembly cycles provides information on the deposition process. The spectra are identical to that observed for PAH--Os in aqueous solution, indicating that the PAH--Os metal complex structure is conserved in the multilayers. We observe at approximately 500 nm incoming and outgoing Raman resonances of osmium and bipyridine vibrational modes. These resonances are associated to the metal-to-ligand charge transfer (MLCT) transition. We study the evolution of these Raman modes as a function of the Os oxidation state during in situ electrochemistry. During the oxidation process, Os(II)-->Os(III), the Raman resonance related to the MLCT disappears and the bipyridine related modes harden by approximately 10 cm(-1). These results are correlated with optical transmission measurements which show the disappearance of the visible region absorption when the Os complex is oxidized. We also find partial quenching of the Raman mode intensity after in situ voltamperometric cycles which demonstrates the existence of photo-electro-chemical processes.

Journal Article↗

Histopathological correlation of cartilage swelling detected by magnetic resonance imaging in early experimental osteoarthritis.

OBJECTIVE: We previously reported that an increase of cartilage thickness is the earliest measurable change by magnetic resonance imaging (MRI) in early stages of experimental osteoarthritis (OA). Our present objective was to study the microscopic translation of this finding in order to know whether the cartilage thickness increment represents the earliest structural damage or whether it alternatively constitutes a non-progressive reversible phenomenon. METHODS: OA was induced by partial medial meniscectomy in rabbits. Normal and sham-operated animals were used as controls. Gross and microscopic cartilage changes were sequentially assessed after surgery at 0, 2, 4, 6, 8, 10 and 52 weeks, and compared to MRI findings. RESULTS: The swelling of cartilage detected by MRI correlated with depletion in matrix proteoglycans and cellular loss, which were closely related to the progression of OA at the earliest stages. Abnormalities of the cartilage structure appeared only in advanced OA. CONCLUSION: Cartilage swelling detected by MRI is due to proteoglycan depletion and represents the earliest abnormality in OA. Because it is accompanied by cellular loss, it cannot be merely attributed to surgical trauma and represents true tissue damage. The biological meaning of volume variations detected by MRI should be assessed carefully taking into account the disease stage as an increase in cartilage height also reflects cartilage damage and not a reparative process.

Animals↗

Reproductive performance of lactating dairy cows treated with cloprostenol at the time of insemination.

The effect of intravenous cloprostenol treatment at the time of insemination on reproductive performance was consecutively evaluated in three different subpopulations of high producing lactating dairy cows: Study (1) early postpartum synchronized and fixed-time inseminated (about 50 days in milk) cows (n = 379: 187 control and 192 treated cows); Study (2) presumed high fertility cows first inseminated between 90 and 120 days postpartum (n = 248: 124 control and 124 treated cows); and Study (3) heat stressed repeat breeder cows (n = 183: 93 control and 90 treated cows). Data were analyzed using multiple regression methods. Study 1: Parity (primiparous versus multiparous), milk production, body condition score at AI, insemination season (cool versus warm period) and treatment were included in the analysis as potential factors affecting ovulation, double ovulation, return to estrus, and pregnancy to first AI and to second AI (first AI plus return AI) rates. Logistic regression analysis indicated that the final model for ovulation rate only included the interaction (P = 0.002) between insemination season and treatment. Cloprostenol treatment at insemination led to a 4.2-fold increase in the ovulation rate in cows inseminated during the warm period. There were no significant effects of treatment, parity, milk production, body score or the insemination season on the return to estrus rate. The only variables included in the final logistic model for double ovulation and pregnancy to first AI rates were treatment and season, respectively. Treatment led to a 2.6-fold increase (P = 0.001) in the double ovulation rate, whereas cows inseminated in the warm period were 2.1 times less likely (P = 0.007) to become pregnant at first AI compared to those inseminated in the cool season. The variables included in the final logistic model for the pregnancy rate to second AI were treatment and season. Cloprostenol given at AI increased the risk of pregnancy 1.9 times (P = 0.002), and cows inseminated during the warm season were two times less likely to become pregnant (P = 0.003). No significant interactions were found among these three dependent variables (double ovulation and pregnancy to first and to second AI rates). Study 2: Logistic regression analysis of all the dependent variables: return to estrus, and pregnancy to first and to second AI (first AI plus return to AI) rates indicated no significant effects of treatment, parity, days in milk, milk production or body score at AI. No significant interactions were found. Study 3: The final model for the pregnancy rate only included the interaction between parity (primiparous versus multiparous) and treatment. Days in milk, milk production and insemination number showed no significant effect on pregnancy rate. Cloprostenol treatment at insemination increased the pregnancy rate in primiparous repeat breeder cows (odds ratio: 3.6). The treatment group and parity showed significant (P < 0.0001) interaction. This interaction suggests that cloprostenol treatment of primiparous cows at insemination might enhance pregnancy yet have no effect in multiparous cows. Our findings indicate that cloprostenol administered at insemination promotes ovulation and double ovulation in lactating dairy cows. Cloprostenol treatment showed no benefit in cows with acceptable reproductive performance, suggesting that cloprostenol treatment at AI may only be useful in cows in which stress factors affect ovulation and in repeat breeder cows.

Animals↗

The transcriptome of adult female Anopheles darlingi salivary glands.

Anopheles (Nyssorhynchus) darlingi is an important malaria vector in South and Central America; however, little is known about molecular aspects of its biology. Genomic and proteomic analyses were performed on the salivary gland products of Anopheles darlingi. A total of 593 randomly selected, salivary gland-derived cDNAs were sequenced and assembled based on their similarities into 288 clusters. The putative translated proteins were classified into three categories: (S) secretory products, (H) housekeeping products and (U) products with unknown cell location and function. Ninety-three clusters encode putative secreted proteins and several of them, such as an anophelin, a thrombin inhibitor, apyrases and several new members of the D7 protein family, were identified as molecules involved in haematophagy. Sugar-feeding related enzymes (alpha-glucosidases and alpha-amylase) also were found among the secreted salivary products. Ninety-nine clusters encode housekeeping proteins associated with energy metabolism, protein synthesis, signal transduction and other cellular functions. Ninety-seven clusters encode proteins with no similarity with known proteins. Comparison of the sequence divergence of the S and H categories of proteins of An. darlingi and An. gambiae revealed that the salivary proteins are less conserved than the housekeeping proteins, and therefore are changing at a faster evolutionary rate. Tabular and supplementary material containing the cDNA sequences and annotations are available at http://www.ncbi.nlm.nih.gov/projects/Mosquito/A_darlingi_sialome/

Animals↗