Neutrophil isolation from feline blood using discontinuous Percoll dilutions

Sürsal, Neslihan; Çakmak, Ayşe; Yıldız, Kader

doi:10.1055/s-0038-1677404

Cited by 1 publication

(7 citation statements)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition, there is no consensus on the best technique for isolating granulocytes to achieve high purity. The methods applied for granulocyte isolation are diverse [29][30][31][33][34][35]53,[67][68][69][70], and their efficiency has been reported in different ways, such as cellular viability or cell isolation yield, cell functionality, number of target cells and contaminants after isolation, which makes it difficult to evaluate the methods for clinical application.…”

Section: Discussionmentioning

confidence: 99%

“…The first stage consisted of the use of Percoll gradients with HBSS (method III), and the second stage used Ficoll on the granulocytes isolated by method III. This first stage of granulocyte isolation was used in some studies as a unique stage of isolation [29,31,32], and the purity analysis, granulocyte yield and cellular viability were reported as 81% purity in 4 × 10 6 cells/mL [31], 95% purity in 2.2 × 10 6 cells/mL [29] and 95% purity for the yield range of 1 to 2 × 10 6 cells/mL, as well as 95% cellular viability [32]. These results corroborate our findings of 86.5% purity in 5.0 × 10 6 cells/mL and 98.3% cellular viability.…”

Section: Discussionmentioning

confidence: 99%

“…Granulocytes can be isolated from whole blood by different methods. The most commonly used method uses gradients of Percoll [29][30][31][32][33][34]. Isolation purity, viability and yield are important factors for evaluating each method and ensuring cellular integrity [29,[34][35][36].The noninvasive evaluation of acute inflammatory processes makes it possible to analyze molecular and cellular processes in vivo, usually based on targeting specialized cells such as macrophages, lymphocytes or granulocytes to the desired target [37].…”

mentioning

confidence: 99%

“…The most commonly used method uses gradients of Percoll [29][30][31][32][33][34]. Isolation purity, viability and yield are important factors for evaluating each method and ensuring cellular integrity [29,[34][35][36].The noninvasive evaluation of acute inflammatory processes makes it possible to analyze molecular and cellular processes in vivo, usually based on targeting specialized cells such as macrophages, lymphocytes or granulocytes to the desired target [37]. Noninvasive imaging techniques that provide structural, functional and molecular information in preclinical and clinical studies include CT, MRI, positron emission tomography (PET), single-photon emission computed tomography (SPETC) and near-infrared fluorescence (NIRF) imaging, among others [38][39][40], each of which has advantages and limitations.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

See 4 more Smart Citations

Methods of Granulocyte Isolation from Human Blood and Labeling with Multimodal Superparamagnetic Iron Oxide Nanoparticles

et al. 2020

View full text Add to dashboard Cite

This in vitro study aimed to find the best method of granulocyte isolation for subsequent labeling with multimodal nanoparticles (magnetic and fluorescent properties) to enable detection by optical and magnetic resonance imaging (MRI) techniques. The granulocytes were obtained from venous blood samples from 12 healthy volunteers. To achieve high purity and yield, four different methods of granulocyte isolation were evaluated. The isolated granulocytes were labeled with multimodal superparamagnetic iron oxide nanoparticles (M-SPIONs) coated with dextran, and the iron load was evaluated qualitatively and quantitatively by MRI, near-infrared fluorescence (NIRF) and inductively coupled plasma mass spectrometry (ICP-MS). The best method of granulocyte isolation was Percoll with Ficoll, which showed 95.92% purity and 94% viability. After labeling with M-SPIONs, the granulocytes showed 98.0% purity with a yield of 3.5 × 10 6 cells/mL and more than 98.6% viability. The iron-loading value in the labeled granulocytes, as obtained by MRI, was 6.40 ± 0.18 pg/cell. Similar values were found with the ICP-MS and NIRF imaging techniques. Therefore, our study shows that it is possible to isolate granulocytes with high purity and yield and labeling with M-SPIONs provides a high internalized iron load and low toxicity to cells. Therefore, these M-SPION-labeled granulocytes could be a promising candidate for future use in inflammation/infection detection by optical and MRI techniques.Hard-to-reach or occult inflammation and infection processes are clinically challenging. Their clinical diagnoses involve biochemical and radiological examinations, but these methods can produce false negatives [11][12][13][14][15]. To determine the localization, extent and severity of inflammation, imaging techniques such as computed tomography (CT) and magnetic resonance imaging (MRI) can be performed but have limitations in the early stages of the disease, as they are dependent on morphological changes [16][17][18]. Accurate and early diagnosis of inflammation and infection helps reduce mortality and morbidity and increase drug treatment success [19][20][21].Proinflammatory effects help leukocytes migrate from the blood vessel to the site of injury, due to increased vascular permeability through stimulation by cytokines and interleukins. There is an initial accumulation of granulocytes, mainly neutrophils, in the inflamed area and later of lymphocytes and macrophages [22][23][24]. Granulocytes are the most abundant subset of leukocytes [25] and are produced in the bone marrow, originating from myeloid precursor cells and undergoing a maturing process before they are able to perform their functions, mainly phagocytosis [25,26]. They differ from other cells in that they have granules with proteolytic enzymes that fight microorganisms or other inflammatory agents [25,27]. Granulocytes are considered frontline cells in inflammation and infection processes [28]. In many cases, to detect inflammation/infection, it is necessary to label isolated granulocyte...

show abstract