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1.
The aim of this study was to validate commercially available methods for porcine haptoglobin (Hp), C-reactive protein (CRP), serum amyloid A (SAA) and major acute phase protein (Pig-MAP) determinations. Intra and inter assay coefficients of variation (CVs) were lower than 20% in all cases with exception of inter assay CVs for CRP and Pig-MAP assays with samples of low acute phase proteins concentration, and for SAA assay at any acute phase proteins concentration. All methods showed good linearity and detection limits were low enough to detect APPs levels in healthy animals. Hp and SAA were very affected by haemolysis. Lipaemia influenced mainly on SAA determination. Over 15-fold increase was observed in CRP and SAA concentrations after artificially induced inflammation by a single subcutaneous dose of turpentine, whereas Hp and Pig-MAP increased less than 5-fold.  相似文献   

2.
The possibility to use acute phase proteins to monitor the elimination of a bacterial infection in pigs would facilitate an objective assessment of treatment with various antimicrobial substances. To examine this possibility, the acute phase response (IL-6, serum amyloid A (SAA), and haptoglobin) elicited by Actinobacillus pleuropneumoniae and its reduction on treatment with various antibiotics was studied in serum from specific pathogen free (SPF) pigs. Pigs were infected intranasally with A. pleuropneumoniae serotype 2, and either left as non-treated control pigs or treated with different antibiotics intramuscularly at onset of respiratory disease (20h post-infection). Pigs responded to the infection with prominent increases in activity and concentrations of IL-6, SAA, and haptoglobin. These responses were to a certain extent overlapping and covered the time span from a few hours after infection until development of detectable levels of specific antibodies (7-10 days post-infection in untreated pigs). The haptoglobin response lasted until the end of the study on day 17 and thereby partly coincided with the antibody response. Treatment with antimicrobials that effectively reduced establishment of the infection with A. pleuropneumoniae also reduced the duration of all three acute phase responses, and reduced the concentration of serum haptoglobin. In contrast, less efficacious treatments did not reduce these acute phase responses. Thus, acute phase reactants can be applied to monitor therapeutic effects of antimicrobial drugs in the pig and measurements of IL-6, SAA and haptoglobin could add valuable information about the stage of infection during a disease outbreak.  相似文献   

3.
In the present study acute phase proteins (APPs) responses in pigs after subclinical infection with H1N1 swine influenza virus (SwH1N1) were evaluated. Fourteen 5 weeks old, seronegative piglets, both sexes were used. Ten of them were infected intranasally with SwH1N1. C-reactive protein (CRP), haptoglobin (Hp), serum amyloid A (SAA) and pig major acute phase protein (Pig-MAP) concentrations in serum were measured using commercial ELISAs. No significant clinical signs were observed in any of the infected pigs, however, all infected animals developed specific antibodies against SwH1N1 and viral shedding was observed from 2 to 5dpi. Only concentrations of Hp and SAA were significantly induced after infection, with mean maximum levels from days 1 to 2 post infection (dpi). The concentrations of CRP and Pig-MAP remained generally unchanged, however in half of infected pigs the concentration of CRP tended to increase at 1dpi (but without statistical significance). The results of our study confirmed that monitoring of APPs may be useful for detection of subclinically infected pigs. The use of SAA or Hp and Pig-MAP may be a valuable in combination [i.e. Hp (increased concentration) and Pig-MAP (unchanged concentration)] to detect subclinically SIV infected pigs, or to identify pigs actually producing a large amount of virus. Additional studies need to be done in order to confirm these findings.  相似文献   

4.
The response of serum amyloid A (SAA) concentrations in serum and saliva during a natural porcine respiratory and reproductive syndrome (PRRS) outbreak was investigated. Basal levels of SAA were reliably determined and significant differences in SAA were found between diseased and healthy pigs in both sample types (serum SAA medians 78.3 vs. 55.74 mg/L, respectively; salivary SAA medians 8.97 vs. 2.52 mg/L, respectively; P < 0.001). In serum, an SAA cut-off value of 66.4 mg/L showed 68.9% sensitivity and 68.09% specificity, while in saliva an SAA cut-off value of 5.59 mg/mL showed a sensitivity–specificity pair of 69–66%. Furthermore, it was observed that the growth stage of animals should be accounted to correctly interpret SAA measurements, since more accurate cut-off values could be determined and a particular behaviour of salivary SAA was identified in post-weaning pigs. Salivary and serum SAA measurements can therefore be confirmed as a valuable tool to consistently discriminate between healthy and PRRS-affected pigs.  相似文献   

5.

Background

An in-clinic assay for equine serum amyloid A (SAA) analysis, Equinostic EVA1, was evaluated for use in a clinical setting. Stability of SAA in serum samples was determined.

Methods

Intra- and inter- assay variation of the in-clinic method was determined. The in-clinic method (EVA1) results were compared to a reference method (Eiken LZ SAA) with 62 patient samples. For samples with SAA concentrations within the assay range of EVA1 (10-270 mg/L), differences between the methods were evaluated in a difference plot. Linearity under dilution was evaluated in two samples. Stability of SAA in three serum pools stored at 4°C and approximately 22°C was evaluated with the reference method day 0, 1, 2, 4, 7, 17 and analysed with a two-way ANOVA.

Results

The imprecision (coefficient of variation, CV) for the in-clinic method was acceptable at higher SAA concentrations with CV values of 7,3-12%, but poor at low SAA concentrations with CV values of 27% and 37% for intra- and inter-assay variation respectively. Recovery after dilution was 50-138%. The in-clinic assay and the reference method identified equally well horses with low (<10 mg/L) and high (>270 mg/L) SAA concentrations. Within the assay range of the in-clinic method, 10-270 mg/L, the difference between the two methods was slightly higher than could be explained by the inherent imprecision of the assays. There were no significant changes of serum SAA concentrations during storage.

Conclusions

The in-clinic assay identified horses with SAA concentrations of <10 mg/L and >270 mg/L in a similar way as the reference method, and provided an estimate of the SAA concentration in the range of 10-270 mg/L. The imprecision of the in-clinic method was acceptable at high SAA concentrations but not at low concentrations. Dilution of samples gave inconsistent results. SAA was stable both at room temperature and refrigerated, and thus samples may be stored before analysis with the reference method.  相似文献   

6.
The diagnostic performance of canine serum amyloid A (SAA) was compared with that of C-reactive protein (CRP) in the detection of systemic inflammation in dogs. Sera from 500 dogs were retrospectively included in the study. C-reactive protein and SAA were measured using validated automated assays. The overlap performance, clinical decision limits, overall diagnostic performance, correlations, and agreement in the clinical classification between these 2 diagnostic markers were compared. Significantly higher concentrations of both proteins were detected in dogs with systemic inflammation (SAA range: 48.75 to > 2700 mg/L; CRP range: 0.4 to 907.4 mg/L) compared to dogs without systemic inflammation (SAA range: 1.06 to 56.4 mg/L; CRP range: 0.07 to 24.7 mg/L). Both proteins were shown to be sensitive and specific markers of systemic inflammation in dogs. Significant correlations and excellent diagnostic agreement were observed between the 2 markers. However, SAA showed a wider range of concentrations and a significantly superior overall diagnostic performance compared with CRP.  相似文献   

7.
Background: Acute phase proteins (APPs) can provide a sensitive test option for detection of inflammation in companion and large animals as well as many nondomesticated mammals. While some basic science studies have addressed APPs expression in rabbits, modern methods have not been widely applied to the detection of inflammation in this species. Methods: In this study, an automated immunoturbidimetric assay for serum amyloid A (SAA) was evaluated. Results: The median (min-max) SAA level of clinically normal rabbits was 6.3 (6.3–24.2) mg/L versus 7.0 (6.3–1388.0) mg/L for the abnormal group. The assay results were also compared to a previously described assay for C-reactive protein (CRP) and found correlated (r = 0.76, P < 0.0001). Additionally, the point-of-care lateral flow device (LFD) for SAA correlated (r = 1.00) when samples were examined as above or below the reference interval determined by the automated SAA assay. Conclusions and clinical relevance: Reference laboratory testing for APPs allows for accurate measurement of SAA and CRP; the LFD offers a new option for point of care testing. Additional studies may show that APPs can have value in the diagnosis and prognosis of diseases in rabbits.  相似文献   

8.
A non-competitive chemiluminescence enzyme immunoassay for measuring serum amyloid A (SAA) in equine serum was developed. A polyclonal anti-equine-amyloid A antiserum specific for equine SAA was utilized, and the assay was standardized using highly purified equine SAA. An acute phase horse serum was calibrated against the purified SAA and was used as standard when running the assay. Serum SAA concentrations in the range of 3-1210 mg/l could be measured. The reference range of SAA in clinically healthy adult horses was <7 mg/l. The clinical validation of the assay comprised the SAA responses after surgery and experimentally induced aseptic arthritis, and those associated with viral and bacterial infections. The SAA response after surgery (castration) was consistent, with peak concentrations on day 2 and a return to normal SAA concentrations within eight days. The aseptic arthritis produced an SAA response with a pattern similar to that seen after surgery, with peak concentrations of SAA 36-48 h after induction. Seven horses showed a biphasic pattern, with a second rise in SAA concentrations on day 4 and 5. All animals had SAA levels <7 mg/l on day 15. All horses with viral and bacterial infections had SAA concentrations above 7 mg/l. The ranges of SAA concentrations following the different types of inflammation overlap, being consistent with the unspecific nature of the SAA response. This study revealed that SAA is a sensitive and unspecific marker for inflammation, and describes the dynamics of the SAA response after standardized and well defined tissue damage.  相似文献   

9.
To establish the relationship between serum levels of three acute phase proteins, haptoglobin (Hp), C-reactive protein (CRP) and serum amyloid A (SAA), and the occurrence and severity of lesions at slaughter, a study was carried out using 70 fattening pigs from a finishing unit. Pigs were divided into three groups: Group 1 (25 pigs with clinical signs of disease), Group 2 (25 apparently healthy pigs with lesions at slaughter) and Group 3 (20 apparently healthy pigs with no lesions at slaughter). Serum levels of CRP, SAA and Hp were significantly higher in pigs with clinical signs of disease than in apparently healthy animals. Additionally, in apparently healthy pigs, serum levels of Hp and CRP were significantly higher in animals with lesions than those without lesions. The extent and severity of lung lesions were related to serum levels of Hp.  相似文献   

10.
This study established the precision and accuracy of a modified latex agglutination turbidimetric immunoassay (LATIA) reagent, and evaluated the ability of the measurement of serum amyloid A (SAA) compared to haptoglobin and α1-acid glycoprotein, which are acute phase proteins (APPs), for diagnosis of clinical mastitis. Concentrations of APPs in cows with mastitis were significantly higher than those in healthy cow. Only the plasma SAA concentration in cows with clinical mastitis (44.90 mg/l; n=15) was significantly higher than that in those with subclinical mastitis (10.70 mg/l; n=16), enabling their diagnosis in contrast to other APPs. Thus, the SAA assay using a LATIA reagent is useful in assessing mastitis severity due to its higher sensitivity and specificity than other APP assays.  相似文献   

11.
Serum amyloid A (SAA) is considered a major acute phase protein (APP) in horses. Serum amyloid A stall-side assays are commercially available to assess the inflammatory response of patients with various infectious and noninfectious conditions. The objective of this study was to determine the analytical performance of a new point-of-care (POC) assay for the measurement of SAA in whole blood and plasma of horses. One hundred and sixty blood samples were collected from 60 horses at various time points after immunization with an equine core vaccine. Analytical validation of the SAA POC assay included the measurement of SAA in whole blood and plasma, assessment of linearity and precision, and comparison of the SAA POC results with those obtained with a validated turbidimetric immunoassay (TIA). The SAA POC assay yielded similar results in whole blood and plasma (P > .05), and the results were positively correlated with the TIA (R2 = 0.964). The assay displayed solid linearity throughout the detection range of ≤ 20 to 3,000 μg/mL (R2 = 0.984) with inter-assay and intra-assay coefficients of variation ranging from 7.8% to 13.3% and 5.7% to 12.0%, respectively. The new SAA POC assay was able to reliably measure SAA in both whole blood and plasma. Similar to previously validated assays, the new SAA POC assay is a valuable tool to investigate the inflammatory response in various clinical diseases of horses.  相似文献   

12.
The aim of this study was to investigate the reliability of an immunoturbidometric assay for measuring the acute phase protein serum amyloid A (SAA) in horses in clinical practice. The assay was compared to a previously validated assay, and overlap performance was assessed by measuring the concentration of SAA in clinically healthy horses and horses with inflammatory and non-inflammatory diseases. In pools of serum with low and high SAA concentrations the assay's intra-assay coefficients of variation were 11.7 per cent and 4.6 per cent, and its interassay coefficients of variation were 9.1 per cent and 5.6 per cent, respectively. Slight inaccuracies were observed, but they were negligible in comparison with the range of the SAA response. The assay systematically underestimated the concentrations of SAA in comparison with the results of the validated assay. The assay detected the expected difference in SAA concentrations between the healthy and diseased horses.  相似文献   

13.
Lipopolysaccharide (LPS) has been widely used as a model of immune challenge in pigs as it induces the immediate synthesis of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and IL-6, which trigger the production of the acute phase proteins (APPs) C-reactive protein (CRP), haptoglobin (Hp) and pig-Major Acute Phase Protein (pig-MAP). To measure secreted proteins in porcine plasma, specific and sensitive Enzyme-Linked Immuno Sorbent Assays (ELISAs) are well-suited to perform single parameter analysis, yet this approach is time-consuming and expensive for multi-parameter analyses. During the last decade, multiplex bead-based flow cytometry has been increasingly applied as it offers the opportunity to estimate protein ratios in a small sample volume. Cytometric bead array (CBA) is a flow cytometric application using a diversity of beads with unique fluorescence intensities, covalently coupled to a capture antibody for each protein of interest. Detection antibodies, either directly or indirectly conjugated to a fluorochrome, are added to accomplish the desired sandwich format. The aim of the present study was to develop a CBA 3-plex assay for the major pro-inflammatory cytokines TNF-α, IL-1β and IL-6, and an additional CBA 2-plex assay for the major APPs, CRP and pig-MAP, in porcine plasma. Results were compared to commercial ELISA kits. For the CBA 3-plex assay, the limits of detection (LODs) varied between 0.005 and 0.363 ng/mL, the intra- and inter-assay coefficients of variation were <10% and <16%, respectively. For TNF-α, IL-1β, IL-6 and pig-MAP, CBA time-concentration profiles similar to those obtained with commercial ELISAs were observed. In conclusion, the novel validated CBA 3-plex assay provides a fast and economical screening tool for determination of pro-inflammatory cytokine profiles in limited porcine plasma volumes. This tool will be applied to study the immunomodulatory properties of drugs in a porcine LPS inflammation model. This study also demonstrated the applicability of CBA for measurement of APPs in pigs, although a different combination than pig-MAP with CRP is recommended.  相似文献   

14.
Intensive exercise results in the increased blood concentration of the acute phase proteins in horses competing in some sport disciplines. In this study, the blood level of serum amyloid A (SAA) was analyzed in Thoroughbred racehorses during 5 days after completion of the race. Samples were collected from 25 healthy Thoroughbred horses beginning with 4 hours after the race and repeated daily up to the fifth day after the race. Serum amyloid A analysis was performed using commercial enzyme-linked immunosorbent assay kit, and the results were presented as median, interquartile range (IQR), and range. Data were analyzed using Friedman's nonparametric analysis of variance. The acute phase response (APR) was reflected by an increased SAA level after the race, reaching significantly higher concentrations on days 1 (P < .001) and 2 (P = .005) and falling below the level of the first sample on day 5 (P = .006). The median peak concentration observed on day 1 after the race was 3.84 mg/L (IQR, 2.32 to 8.86). Racing induces minute changes in SAA concentration typical for the exercise-induced APR; however, the significance of this reaction in the context of horse health and fitness remains unclear.  相似文献   

15.
The pig acute phase protein (APP) response to experimental Streptococcus suis (S. suis) infection was mapped by the measurement of the positive APPs C-reactive protein (CRP), serum amyloid A (SAA), haptoglobin (Hp) and major acute phase protein (pig-MAP) and the negative APPs albumin and apolipoprotein (Apo) A-I. The aim was to elucidate the differences in the acute phase behaviour of the individual APPs during a typical bacterial septicaemic infection. Pigs were inoculated subcutaneously with live S. suis serotype 2 and blood was sampled before and on various days post inoculation (p.i.), until the pigs were killed and autopsied on day 14 p.i. Clinical signs (fever and lameness) were observed in four of the five inoculated pigs from day 2 p.i., and these pigs also had arthritic lesions at autopsy. CRP and SAA showed fast increases in serum concentrations, CRP being elevated from days 1 to 12 p.i. and peaking at 10 times the day 0-levels on day 1 p.i. SAA rose quickly to peak levels of 30-40 times the day 0-level on days 1-2 and returned to pre-inoculation level on day 5 p.i. Hp and pig-MAP showed slightly slower responses, both peaking around 5 days p.i. Hp was increased throughout the experiment with maximum levels around 10 times the day 0-levels, and pig-MAP was elevated on days 1-12 p.i. with peak levels of around seven times the day 0-levels. Apo A-I was decreased from days 1 to 8 and showed minimum levels of about 40% of day 0-levels around 1-2 days p.i. No clear pattern of changes in albumin levels could be identified. One pig, showing clinical signs on day 2 only, also showed an APP response, although of a relatively short duration, whereas three pigs presenting clinical signs for several days had a more protracted acute phase response. Remarkably, the one pig showing no clinical signs and no arthritic lesions showed an APP response comparable to that of the other, clinically affected pigs. Thus, both acute clinical and subclinical S. suis infection could be revealed by the measurement of one or more of the APPs CRP, SAA, Hp, pig-MAP and Apo A-I. The combined measurement of two or three APPs, including proteins with slow and fast kinetics, should be used to achieve the highest sensitivity for the detection of ongoing S. suis infection during a prolonged time period. A diagnostic tool based on such APP-measurements could considerably improve strategic control procedures for this important infection.  相似文献   

16.
Five acute phase proteins (APPs) [C-reactive protein (CRP), serum amyloid A (SAA), haptoglobin (Hp), pig-MAP and albumin] were measured in pigs with naturally occurring infections by porcine reproductive and respiratory syndrome virus (PRRSV), Aujeszky's disease virus (ADV), porcine circovirus type 2 (PCV2) and Mycoplasma hyopneumoniae, and in animals with tail and ear bites, arthritis and other acute inflammatory processes. Healthy specific pathogen-free (SPF) pigs were used as controls. In PRRSV-infected pigs, all APPs with the exception of pig-MAP exhibited significant changes compared with controls. In animals affected with ADV only Hp presented changes of statistical significance, whereas pigs with PCV2 showed marked modifications in all APPs tested. Animals affected with Mycoplasmosis showed concentrations of all positive APPs significantly above levels obtained in SPF pigs, though albumin concentrations did not differ from controls. Finally, all APPs studied showed substantial changes in pigs with acute inflammation. The results indicated that an acute phase response was developed in the different diseases studied, this response being higher in animals with clinical signs and concurrent bacterial processes. Haptoglobin would be the APP that better reflects pathological states; however, to get more complete and valuable information it might be advisable to perform APPs profiles including another APP, such as CRP or SAA.  相似文献   

17.
A specific enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to the porcine pathogen Lawsonia intracellularis was developed and evaluated using sera from na?ve, naturally infected as well as experimentally infected pigs. On the basis of 37 serum samples collected from experimentally infected pigs and 62 serum samples from naturally infected pigs the sensitivity of the ELISA was calculated to 98.0%. The specificity of the test was 99.3%, calculated on the basis of 273 serum samples collected in six herds free of L. intracellularis after medicated eradication. The novel ELISA was a specific and sensitive method for detecting specific antibodies, and may be a good alternative to the existing serological tests for L. intracellularis. It may be usable for diagnosis of proliferative enteropathy and for determination of a herd's epidemiologic status.  相似文献   

18.
The objective of this study was to determine the serum concentration levels of selected acute phase proteins (APP), haptoglobin (HPT) and pig-major acute phase protein (pig-MAP), in postweaning multisystemic wasting syndrome (PMWS) affected pigs and PCV2-subclinically infected pigs. In a first study, a group of 15 eight-week-old conventional pigs from a PMWS affected farm were bled and a complete necropsy, histopathology and in situ hybridisation to detect PCV2 were performed. Based on the results, pigs were classified as suffering from PMWS (n = 10) or healthy animals (n = 5). In a second study, a group of 45 pigs from another PMWS affected farm were selected and bled at 3, 7, 12 and 28 weeks of age. The assessment of PCV2 infection status in these pigs was retrospectively done by PCV2 PCR in serum samples. Selected APP were measured in the serum of all studied pigs by means of radial immunodiffusion. Mean HPT and pig-MAP levels were significantly increased (p = 0.004 and p = 0.0006 respectively) in PMWS-affected pigs when compared to levels found in healthy pigs (2.52 +/- 0.88 mg/mL vs. 1.06 +/- 0.73 mg/mL for HPT and 3.81 +/- 1.53 mg/mL vs. 0.76 +/- 0.34 mg/mL for pig-MAP). In the second study, no significant difference in mean HPT and pig-MAP values were observed between PCV2 PCR positive and negative pigs of any age. However, both APP increased significantly with age in PCV2 PCR negative pigs. Altogether, the present results suggest that APP levels are significantly increased in pigs that develop PMWS, but not in animals with a PCV2 subclinical infection.  相似文献   

19.
A competitive enzyme-linked immunosorbent assay (ELISA) for the detection of swine trichinosis has been developed using a biotinylated monoclonal antibody and an avidin-enzyme conjugate. The assay is based on competitive binding between swine serum antibodies and a monoclonal antibody specific for an antigenic determinant present on proteins from Trichinella spiralis excretory-secretory products with molecular weights of 45,000, 49,000, and 53,000. The competitive ELISA reliably detected pigs infected experimentally with T. spiralis and eliminated false-positive reactions in pigs infected with other swine nematodes, particularly Trichurus suis. When the competitive ELISA and an indirect ELISA using affinity-isolated antigen were compared using serum from pigs with naturally-acquired infections of T. spiralis, both tests were highly effective in detecting infected animals.  相似文献   

20.
New tools are needed to detect chronic sub-clinical mastitis, especially in automatic milking systems. Haptoglobin and serum amyloid A (SAA) are the two most sensitive bovine acute phase proteins, and their concentrations increase in milk from cows with clinical mastitis and in milk from cows with experimentally induced chronic sub-clinical Staphylococcus aureus mastitis. The aim of this study was to further evaluate the potential for haptoglobin and SAA in milk as indicators of chronic sub-clinical mastitis. Quarter milk samples were collected from 41 cows with a mean composite milk somatic cell count (CSCC) above 300,000 cells/mL during at least two months prior to sampling. Quarter milk samples were also taken from eleven cows with a mean CSCC below 80,000 cells/mL during at least two previous months. These samples were analysed for haptoglobin, SAA, adenosine triphosphate (ATP) activity and bacterial growth. The samples were grouped according to their ATP, haptoglobin and SAA status. ATP+ samples had ATP > 2 x 10(-10) mol/mL, Hp+ and SAA+ samples had detectable levels of haptoglobin (> or = 0.3 mg/L) and SAA (> or = 0.9 mg/L), respectively. In udder quarter samples from healthy cows, 42 out of 44 samples belonged to the ATP-Hp-SAA- group. Among cows with chronic sub-clinical mastitis, the ATP+Hp+SAA+ group contained 66 out of 164 samples while 44 samples belonged to the ATP+Hp-SAA- group. Detectable levels of haptoglobin and SAA were found in 92 and 80 samples, respectively. Growth of udder pathogens was detected in 28 samples and Staphylococcus aureus was the most common bacteria. In conclusion, haptoglobin and SAA concentrations below the detection limit were considered as good indicators of healthy udder quarters. A substantial variation in haptoglobin and SAA concentrations in milk was observed in udder quarters with chronic sub-clinical mastitis.  相似文献   

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