Revisiting the dilution factor as vital parameter for sensitivity of ELISA assay in CSF and Plasma

Thakur, Keshav; Sharma, Suresh; Prabhakar, Sudesh; Gupta, Pawan Kumar; Anand, Akshay

doi:10.5214/ans.0972.7531.220108

Cited by 5 publications

(6 citation statements)

References 30 publications

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“…Although this kit was used successfully for studies in cats, whether the response level (cross reactivity) was similar for human and for feline samples remains to be confirmed by the manufacturer. One possible explanation for the difference in the response level is the sample dilution effect (in which after the response goes beyond the linear range of the assay, the sample must be further diluted) and might start to exhibit a disproportionate increase in response . In addition, our study focused on the difference between the progression and non‐progression groups, in which some data came from younger and less severe CKD cats that could have brought the median FGF‐23 concentration lower and mostly within the assay, linear range.…”

Section: Discussionmentioning

confidence: 99%

The association of indoxyl sulfate with fibroblast growth factor‐23 in cats with chronic kidney disease

Liao

Chou

Lee

2019

Veterinary Internal Medicne

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Background Indoxyl sulfate (IS) has been reported not only to increase with the severity of impaired renal function, but also possibly to be a factor associated with bone abnormalities linked to fibroblast growth factor‐23 (FGF‐23) in humans with chronic kidney disease (CKD). It is not yet known whether this correlation between IS and FGF‐23 holds true for cats with CKD. Hypothesis Accumulation of IS is related to FGF‐23 secretion in cats with CKD. Animals Twenty clinically healthy cats and 73 cats with CKD cases were evaluated retrospectively. Methods The concentrations of IS and FGF‐23 in plasma were determined by high‐performance liquid chromatography and ELISA, respectively. Progression was defined as an increment of 0.5 mg/dL of serum creatinine concentration within 3 months. Results Plasma IS and FGF‐23 concentrations were significantly increased concurrently with decreasing renal function. Higher concentration of FGF‐23 was significantly associated with higher concentration of IS after adjusting for various confounding factors including creatinine and phosphate. Furthermore, the correlation between IS and phosphate was higher than that between FGF‐23 and phosphate. When the renal progression group was compared with the non‐progression group, both IS and FGF‐23 were found to be significantly increased (P < .05). In addition, the area under receiver operator curve of the combination of IS and FGF‐23 predicted renal progression at a level >0.9. Conclusions and Clinical Importance Both FGF‐23 and IS are associated with phosphate metabolism and CKD progression.

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Section: Discussionmentioning

confidence: 99%

The association of indoxyl sulfate with fibroblast growth factor‐23 in cats with chronic kidney disease

Liao

Chou

Lee

2019

Veterinary Internal Medicne

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“…On the basis of optimized dilution factors, major allergen proteins were estimated in peanut seeds with increased precision. The results from our study show the importance of standard curve and dilution as an important parameter for quantifying the ELISA results (39). Dilution factor is an important step in ELISA before analyzing the results.…”

Section: Discussionmentioning

confidence: 86%

“…Dilution assay apprises the actual concentration would be taken to experiment. To attained highly sensitive results for protein concentration, optimum values for dilution assay would provide a valid conclusion (39, 40). Earlier few attempts were made to quantify the levels of allergens in peanut seed using immuno assays and chromatography methods.…”

Section: Discussionmentioning

confidence: 99%

An Improved Enzyme-Linked Immunosorbent Assay (ELISA) Based Protocol Using Seeds for Detection of Five Major Peanut Allergens Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8

et al. 2019

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Peanut allergy is an important health concern among many individuals. As there is no effective treatment to peanut allergy, continuous monitoring of peanut-based products, and their sources is essential. Precise detection of peanut allergens is key for identification and development of improved peanut varieties with minimum or no allergens in addition to estimating the levels in peanut-based products available in food chain. The antibody based ELISA protocol along with sample preparation was standardized for Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8 to estimate their quantities in peanut seeds. Three different dilutions were optimized to precisely quantify target allergen proteins in peanut seeds such as Ara h 1 (1/1,000, 1/2,000, and 1/4,000), Ara h 2 and Ara h 3 (1/5,000, 1/10,000, and 1/20,000), Ara h 6 (1/40,000, 1/80,000, and 1/1,60,000), and Ara h 8 (1/10, 1/20, and 1/40). These dilutions were finalized for each allergen based on the accuracy of detection by achieving <20% coefficient of variation in three technical replicates. This protocol captured wide variation of allergen proteins in selected peanut genotypes for Ara h 1 (77–46,106 μg/g), Ara h 2 (265–5,426 μg/g), Ara h 3 (382–12,676 μg/g), Ara h 6 (949–43,375 μg/g), and Ara h 8 (0.385–6 μg/g). The assay is sensitive and reliable in precise detection of five major peanut allergens in seeds. Deployment of such protocol allows screening of large scale germplasm and breeding lines while developing peanut varieties with minimum allergenicity to ensure food safety.

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“…In conditions at which there was a problem in the assay like giving an error as the assay range is not within the detection range of the kit, dilution of the sample done, and then reassessment done. The value of reassessment is then multiplied by the percent of diluting fluid used giving us the actual level which is higher than the detection range [12].…”

Section: Methodsmentioning

confidence: 99%

Apolipoproteins A1 and B and their ratio in acute ischemic stroke patients with intracranial and extracranial arterial stenosis: an Egyptian study

Fahmy

Awady

Sharaf

et al. 2020

Egypt J Neurol Psychiatry Neurosurg

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Background Apolipoprotein A1 and B and their ratio are considered a better biomarker for cardiovascular diseases than a lipid profile, but this previous finding is not proved to cerebrovascular ischemic diseases. The aim of this study is to assess the relation between apolipoprotien A1 and B and their ratio to intra- and extracranial carotid atherosclerosis in patients with ischemic acute stroke. Methods 90 Egyptian patients with acute ischemic stroke are included in the study, and they have been classified into 3 groups: group 1 includes 30 patients with intracranial stenosis, group 2 includes 30 patients with extracranial arterial stenosis, and group 3 includes 30 patients with non-arterial stenosis. Patients were subjected to clinical assessment, routine laboratory measures, and Color-Coded Duplex Sonography for extracranial and intracranial arteries. The measurement of serum levels of apolipoproteins A1 and B was done using enzyme-linked immuno-sorbent assay (ELISA). Results A statistically significant difference was found between patient groups as regards the frequency of abnormal serum LDL-cholesterol (p = 0.04), being elevated in patients with extracranial stenosis (p = 0.01). There was a significant difference between patients groups as regards the frequency of abnormal serum HDL-cholesterol (p = 0.02), being lower in patients with extracranial stenosis. High Apo B/A1 ratio was an independent risk factor for intracranial arterial stenosis (p = 0.045). An abnormal elevation of serum LDL cholesterol was an independent risk factor of extracranial arterial stenosis (p = 0.021). Conclusion Apo B/A1 ratio is an independent risk factor for intracranial arterial stenosis, while serum LDL cholesterol is an independent risk factor for extracranial arterial stenosis. Apo B/A1 ratio and serum LDL cholesterol are reliable serum biomarkers for cranial arterial stenosis in acute ischemic stroke patients.

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Revisiting the dilution factor as vital parameter for sensitivity of ELISA assay in CSF and Plasma

Cited by 5 publications

References 30 publications

The association of indoxyl sulfate with fibroblast growth factor‐23 in cats with chronic kidney disease

The association of indoxyl sulfate with fibroblast growth factor‐23 in cats with chronic kidney disease

An Improved Enzyme-Linked Immunosorbent Assay (ELISA) Based Protocol Using Seeds for Detection of Five Major Peanut Allergens Ara h 1, Ara h 2, Ara h 3, Ara h 6, and Ara h 8

Apolipoproteins A1 and B and their ratio in acute ischemic stroke patients with intracranial and extracranial arterial stenosis: an Egyptian study

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