· Functional test ex vivo

來自體內(nèi)的功能測試

· Evaluation of single cells to detect heterogenous populations

評價檢測單個細胞異質(zhì)種群

· Whole blood assay: No isolation procedures and optimal culture medium

全血檢測：沒有隔離程序和*培養(yǎng)基

· Physiological stimulans for phagocytes: Bacteria and fMLP

吞噬細胞生理興奮勁：細菌和fMLP

· Dose response: Low and high stimulant

沒有靜電工件比較乳膠珠子聚苯乙烯管內(nèi)

· Standardized test procedure

標準化測試程序

· Exclusion of aggregation artifacts by DNA staining

通過DNA染色排除聚合工件

· Compatible with whole blood of mice and rats

兼容的小鼠和大鼠全血

· Fast assay: Whole assay time is 1.5 hours

快速測定:整個試驗時間是1.5小時

100 analyses. Clinical Diagnostic for the Quantitative Analysis of Leukocyte Oxidative Burst in Human Whole Blood. Evaluation by flow cytometry.

100次分析。臨床診斷人體全血白細胞氧化破裂的定量檢測。通過流式細胞術(shù)進行評估。

Clinical Diagnostic for the Quantitative Analysis of Leukocyte Oxidative Burst in Human Whole Bloods

SUMMARY and EXPLANATION

BURSTTEST (PHAGOBURST) allows the quantitative determination of leukocyte oxidative burst in heparinized whole blood. It contains unlabeled opsonized bacteria (E.coli), phorbol 12-myristate 13-acetate (PMA) and the chemotactic peptide N-formyl-Met-Leu-Phe (fMLP) as stimulants, Dihydrorhodamine (DHR) 123 as a fluorogenic substrate and necessary reagents. It determines the percentage of phagocytic cells which produce reactive oxidants (conversion of DHR 123 to R 123) and their enzymatic activity (amount of R 123 per cell).

The evaluation of these bioactivities should be performed by flow cytometry.

BURSTTEST(PHAGOBURST)允許定量測定白細胞氧化闖入肝素化全血。 它包含未標記的促進調(diào)理作用的細菌(大腸桿菌),佛波醇12十四烷酸乙酸13(PMA)和趨化作用的多肽n甲酰遇見亮氨酸板式換熱器(fMLP),二氫若丹明(DHR)123作為一個熒光襯底和必要的試劑。它決定了吞噬細胞產(chǎn)生活性氧化劑的比例(轉(zhuǎn)換DHR 123 到 R 1233)及其酶活性(每個細胞R 123的數(shù)量)。

這些生物活性的測定應(yīng)該有流式細胞術(shù)完成。

APPLICATIONS

The diagnostic kit is intended to investigate the altered oxidative burst activity found in various disorders and to evaluate the effects of drugs.

Reduced or missing burst activity is observed in inborne defects like the chronic granulomatous disease (CGD). CGD is a heterogenous group of inherited disorders that usually manifests itself during the first two years of life (3, 4). The disease is characterized clinically by repeated and life-threatening infections caused by bacterial and fungal organisms. These infections typically consist of pneumonia, lymphadenitis, or abscesses that involve lymph nodes, lungs, and liver. The NADPH oxidase is the enzyme system responsible for producing superoxide anion, which is quickly converted to hydrogen peroxide and hydroxyl radicals. Abnormalities in the constituent peptides of the NADPH oxidase enzyme system lead to the dysfunctions characteristic of CGD. Neutrophils from CGD patients fail to produce a significant oxidative burst following stimulation. Different forms of CGD are described (classical X-linked CGD and autosomal recessive patterns). BURSTTEST (PHAGOBURST?) is a rapid and sensitive method for the diagnosis of CGD and for the detection of X-linked carriers.

The oxidative burst of granulocytes is impaired in transplant patients and patients with AIDS (6). The spontaneous and fMLP-induced neutrophil respiratory burst was shown to be increased in neonates with laboratory signs of infection (7). Various immunomodulators (e.g., cytokines (GM-CSF, G-CSF, TNF) or drugs) seem to have effects on the oxidative burst. By using fMLP as a low stimulant one can investigate additive or priming effects (8) of test substances.

The diagnostic kit is also applicable on blood of mice, rats, rabbits, dogs, cattle and other species.

診斷測試組旨在研究改變了的氧化破裂活動中發(fā)現(xiàn)各種障礙和評估藥物的影響。

觀察細胞的減少或缺失的破裂活動像慢性肉芽腫性疾病(CGD)

慢性肉芽腫性疾病是一個異構(gòu)群遺傳疾病,通常體現(xiàn)在生命的zui初兩年期間(3、4)。這種疾病的臨床特征是由于細菌和真菌重復(fù)和危機生命的感染。這些感染通常包括肺炎、淋巴腺炎或膿腫涉及到淋巴結(jié)、肺和肝。NADPH氧化酶的酶系統(tǒng)是負責生產(chǎn)超氧化物陰離子,迅速轉(zhuǎn)化為過氧化氫和羥基自由基。NADPH氧化酶系統(tǒng)在組成多肽中的畸形導(dǎo)致慢性肉芽腫性疾病功能障礙性特點。CGD患者的中性粒細胞刺激后無法產(chǎn)生顯著的氧化破裂。BURSTTEST (PHAGOBURST?)是一種快速有效診斷慢性肉芽腫性疾病和檢測X連鎖隱性遺傳疾病攜帶者的方法。

氧化破裂的粒細胞在需要移植的病人和艾滋病患者中受損。無意識和誘發(fā)性fMLP嗜中性粒細胞破裂被證明會增加新生兒實驗室感染的跡象。各種免疫調(diào)節(jié)劑（如細胞激素(GM-CSF, G-CSF, TNF)或藥品）似乎會對氧化破裂產(chǎn)生影響。通過使用fMLP可以研究添加劑或激發(fā)效應(yīng)(8)的測試物質(zhì)。

診斷試劑盒也適用于小老鼠、大老鼠、兔子、狗、牛和其他物種的血液。

TEST PRINCIPLES
Phagocytosis by polymorphonuclear neutrophils and monocytes constitutes an essential arm of host defense against bacterial or fungal infections. The phagocytic process can be separated into several major stages: chemotaxis (migration of phagocytes to inflammatory sites), attachment of particles to the cell surface of phagocytes, ingestion (phagocytosis) and intracellular killing by oxygen-dependent (oxidative burst) and oxygen-independent mechanisms (1, 2).

BURSTTEST (PHAGOBURST?) allows the quantitative determination of leukocyte oxidative burst. The BURSTEST kit contains unlabelled opsonized E.coli bacteria as particulate stimulus, the protein kinase C ligand phorbol 12-myristate 13-acetate (PMA) as high stimulus and the chemotactic peptide N-formyl-MetLeuPhe (fMLP) as low physiological stimulus, Dihydrorhodamine (DHR) 123 as a fluorogenic substrate (5) and necessary reagents. Heparinized whole blood is incubated with the various stimuli at 37°C, a sample without stimulus serves as negative background control. Upon stimulation, granulocytes and monocytes produce reactive oxygen metabolites (superoxide anion, hydrogen peroxide, hypochlorous acid) which destroy bacteria inside the phagosome. Formation of the reactive oxidants during the oxidative burst can be monitored by the addition and oxidation of DHR 123. The reaction is stopped by addition of LYSING SOLUTION, which removes erythrocytes and results in a partial fixation of leukocytes. After one washing step with WASHING SOLUTION, DNA STAINING SOLUTION is added to exclude aggregation artifacts of bacteria or cells. The percentage of cells having produced reactive oxygen radicals are then analyzed as well as their mean fluorescence intensity (enzymatic activity).

溫馨提示：不可用于臨床治療。