This kit can be used in quantitative and qualitative analysis of gentamicin residue in vaccine and cell culture.
Contents of Kit
No.
Components
Size
Storage Conditions
1
Microtiter plate
96 wells
2-8°C
2
Standard solutions
6×1ml/bottle
2-8°C
3
High concentration Gentamicin Standard 1μg/ml
1ml/bottle
2-8°C
4
Enzyme conjugate
12ml
2-8°C
5
Antibody solution
7ml
2-8°C
6
Substrate Solution
2×7ml
2-8°C
7
Stop solution
7ml
2-8°C
8
20×concentrated wash solution
40ml
2-8°C
9
2×Sample diluent
50ml
2-8°C
Storage
2-8°C, 12 months.
Precision
Intra-assay and inter-assay of the ELISA kit is less than 5%
Detection Range
0.1-8.1ng/mL
Sensitivity
0.1ng/mL
General Description
Gentamicin residue in the production of biological samples may lead to severe allergic reactions in certain groups. Thus it is strictly controlled in many countries in the world. This kit is a new product for drug residual detection based on ELISA technology, which is rapid, easy-to-use, and sensitive, and can considerably minimize operation errors and work intensity.
Citations
Publication ()
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The antimalarial drug mefloquine enhances TP53 premature termination codon readthrough by aminoglycoside G418
Michael W Ferguson, Chloe A N Gerak, Christalle C T Chow, Ettore J Rastelli, Kyle E Elmore, Florian Stahl, Sara Hosseini-Farahabadi, Alireza Baradaran-Heravi, Don M Coltart, Michel Roberge
Applications: ELISA Reactive species: Unspecified reactive species
"Abstract: Nonsense mutations constitute ~10% of TP53 mutations in cancer. They introduce a premature termination codon that gives rise to truncated p53 protein with impaired function. The aminoglycoside G418 can induce TP53 premature termination codon readthrough and thus increase cellular levels of full-length protein. Small molecule phthalimide derivatives that can enhance the readthrough activity of G418 have also been described. To determine whether readthrough enhancers exist among drugs that are already approved for use in humans, we tested seven antimalarial drugs for readthrough of the common R213X TP53 nonsense mutation in HDQ-P1 breast cancer cells. Mefloquine induced no TP53 readthrough activity as a single agent but it strongly potentiated readthrough by G418. The two enantiomers composing pharmaceutical mefloquine potentiated readthrough to similar levels in HDQ-P1 cells and also in SW900, NCI-H1688 and HCC1937 cancer cells with different TP53 nonsense mutations. Exposure to G418 and mefloquine increased p53 phosphorylation at Ser15 and P21 transcript levels following DNA damage, indicating p53 produced via readthrough was functional. Mefloquine does not appear to enhance readthrough via lysosomotropic effects as it did not significantly affect lysosomal pH, the cellular levels of G418 or its distribution in organellar or cytosolic fractions. The availability of a readthrough enhancer that is already approved for use in humans should facilitate study of the therapeutic potential of TP53 readthrough in preclinical cancer models." Article snippet: G418 levels were determined using an indirect competitive gentamicin ELISA kit (Creative Diagnostics, DEIA047), according to the manufacturer’s protocol, except that standard curves were generated using G418 diluted in NP-40 lysis buffer or in digitonin lysis buffer.
Figure 1. Detection of G418 accumulation in cells using an indirect competitive gentamicin assay kit
Background
Gentamicin is a highly effective aminoglycoside antibiotic primarily used to treat common infections caused by aerobic Gram-negative bacteria. Its bactericidal activity makes it a favorable option for treating various types of bacterial infections such as pelvic inflammatory disease, pneumonia, bone infections, endocarditis, meningitis, sepsis, and urinary incontinence intestinal infections. Administration of gentamicin is usually by parenteral route, including systemic, topical, and ophthalmic formulations, because its gastrointestinal absorption is minimal.
Figure 1. Structure of Gentamicin. (Source: Naveed, S. et al., 2014)
In addition to clinical applications, gentamicin is a very thermostable antibiotic that remains active even after autoclaving, which makes it widely used in culture media formulations. However, gentamicin is neurotoxic, nephrotoxic, and ototoxic. Its residues in animal food and biological drugs can affect human health and even cause allergic reactions. Therefore, it is strictly controlled in many countries around the world. This kit uses an indirect competitive ELISA method to detect the total amount of gentamicin in cells, tissues, serum, and other samples. Compared with traditional methods, the Gentamicin ELISA kit is simple, fast, sensitive, and suitable for the detection of large quantities of samples.
Q: Our samples derived from cell culture and virus culture. Can I use this kit?
A: This kit can be used in quantitative and qualitative analysis of gentamicin residue in vaccine and cell culture.
Q: Has this kit ever been tested against samples containing G418 (Geneticin) instead of Gentamicin?
A: This kit has not been tested against samples containing G418 (Geneticin) in our lab
Q: Whether I could dilute out the High Concentration Gentamicin Standard to a range of say 0, 0.25, 0.50, 0.75, and 1.0 ug/ml and use this as my standard curve instead?
A: High Concentration Gentamicin Standard are mainly used for more diluted standards. You'd better use them to find the best dilution factor of your sample in according with our typical curve.
Q: Whether the kit will work for higher concentrations of Gentamicin in my samples as well as using this higher concentration standard curve?
A: The concentration of sample in the test beyond the cure of DEIA042 will turn out to be inaccurate.
Q: I've been reading the protocol for the Gentamicin ELISA kit and am curious what the 1 ug/mL gentamicin standard is for?
A: It is used for spike controls.
Q: What kind of samples has this kit been tested with?
A: We have tested CHO cells, HEK293 cell supernatant, hay fermentation broth, vaccine supernatant
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References
An integrative overview on the mechanisms underlying the renal tubular cytotoxicity of gentamicin
Toxicological Sciences
Authors: Quiros Y, Vicente-Vicente L, Morales A I, et al.
Gentamicin is an aminoglycoside antibiotic widely used against infections by Gram-negative microorganisms. Nephrotoxicity is the main limitation to its therapeutic efficacy. Gentamicin nephrotoxicity occurs in 10–20% of therapeutic regimes. A central aspect of gentamicin nephrotoxicity is its tubular effect, which may range from a mere loss of the brush border in epithelial cells to an overt tubular necrosis. Tubular cytotoxicity is the consequence of many interconnected actions, triggered by drug accumulation in epithelial tubular cells. Accumulation results from the presence of the endocytic receptor complex formed by megalin and cubulin, which transports proteins and organic cations inside the cells. Gentamicin then accesses and accumulates in the endosomal compartment, the Golgi and endoplasmic reticulum (ER), causes ER stress, and unleashes the unfolded protein response. An excessive concentration of the drug over an undetermined threshold destabilizes intracellular membranes and the drug redistributes through the cytosol. It then acts on mitochondria to unleash the intrinsic pathway of apoptosis. In addition, lysosomal cathepsins lose confinement and, depending on their new cytosolic concentration, they contribute to the activation of apoptosis or produce a massive proteolysis. However, other effects of gentamicin have also been linked to cell death, such as phospholipidosis, oxidative stress, extracellular calcium–sensing receptor stimulation, and energetic catastrophe. Besides, indirect effects of gentamicin, such as reduced renal blood flow and inflammation, may also contribute or amplify its cytotoxicity. The purpose of this review was to critically integrate all these effects and discuss their relative contribution to tubular cell death.
Gentamicin vestibulotoxicity
Otolaryngology—Head and Neck Surgery
Authors: Halmagyi G M, Fattore C M, Curthoys I S, et al.
We reviewed 36 patients with gentamicin vestibulotoxicity to determine its relationship to gentamicin dosage, serum gentamicin levels, and the development of gentamicin nephrotoxicity. Thirty of the patients had received intravenous or intramuscular gentamicin; six had received intraperitoneal gentamicin. Sixteen of the 30 patients treated with intramuscular or intravenous gentamicin had received less than the recommended maximum dose of 5 mg/kg/day for less than the recommended maximum period of 10 days. Nephrotoxicity as well as vestibulotoxicity developed in 16 of these 30 patients. Gentamicin vestibulotoxicity was not recognized before discharge from hospital in 32 of the 36 patients. We conclude that as far as the vestibular system is concerned there is no safe gentamicin dose and no safe serum gentamicin level, and there is an increased risk of vestibulotoxicity in patients in whom nephrotoxicity develops. Physicians who use gentamicin should become more aware of the clinical features of vestibulotoxicity because stopping gentamicin as soon as symptoms of vestibulotoxicity appear could prevent permanent impairment of vestibular function.
Theophylline dosing and pharmacokinetics for renal protection in neonates with hypoxic-ischemic encephalopathy undergoing therapeutic hypothermia
PEDIATRIC RESEARCH
Authors: Frymoyer, Adam; Van Meurs, Krisa P.; Drover, David R.; Klawitter, Jelena; Christians, Uwe; Chock, Valerie Y.
Background Theophylline, a non-selective adenosine receptor antagonist, improves renal perfusion in the setting of hypoxia-ischemia and may offer therapeutic benefit in neonates with hypoxic-ischemic encephalopathy (HIE) undergoing hypothermia. We evaluated the pharmacokinetics and dose-exposure relationships of theophylline in this population to guide dosing strategies. Methods A population pharmacokinetic analysis was performed in 22 neonates with HIE undergoing hypothermia who were part of a prospective study or retrospective chart review. Aminophylline (intravenous salt form of theophylline) was given per institutional standard of care for low urine output and/or rising serum creatinine (5 mg/kg intravenous (i.v.) load then 1.8 mg/kg i.v. q6h). The ability of different dosing regimens to achieve target concentrations (4-10 mg/L) associated with clinical response was examined. Results Birth weight was a significant predictor of theophylline clearance and volume of distribution (p < 0.05). The median half-life was 39.5 h (range 27.2-50.4). An aminophylline loading dose of 7 mg/kg followed by 1.6 mg/kg q12h was predicted to achieve target concentrations in 84% of simulated neonates. Conclusions In neonates with HIE undergoing hypothermia, theophylline clearance was low with a 50% longer half-life compared to full-term normothermic neonates without HIE. Dosing strategies need to consider the unique pharmacokinetic needs of this population. Impact Theophylline is a potential renal-protective therapy in neonates with HIE undergoing therapeutic hypothermia; however, the pharmacokinetics and dose needs in this population are not known. Theophylline clearance was low in neonates with HIE undergoing therapeutic hypothermia with a 50% longer half-life compared to full-term normothermic neonates without HIE. As theophylline is advanced in clinical development, dosing strategies will need to consider the unique pharmacokinetic needs of neonates with HIE undergoing therapeutic hypothermia.
Umbelliferone attenuates gentamicin-induced renal toxicity by suppression of TLR-4/NF-kappa B-p65/NLRP-3 and JAK1/STAT-3 signaling pathways
ENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH
Authors: Hassanein, Emad H. M.; Ali, Fares E. M.; Kozman, Magy R.; Abd El-Ghafar, Omnia A. M.
Nephrotoxicity is the most common adverse effect of gentamicin (GNT). This study aimed to investigate the possible nephroprotective effect of umbelliferone (UMB), against GNT-induced nephrotoxicity. Rats were allocated into the control group; UMB group (50 mg/kg/day, P.O. for 15 days); GNT group (100 mg/kg/day, i.p., for 8 days); and GNT + UMB group. By the end of the experimental period, serum creatinine, urea, and uric acid as well as urine KIM-1 and urine albumin/creatinine ratio were evaluated to estimate kidney function. Moreover, tissue samples were collected for assessment of ERK1/2, p-ERK1/2, TLR-4, p38 MAPK, NF-kappa B-p65, NLRP-3, IkB alpha, TNF-alpha, IL-1 beta, JAK1, STAT-3, p-STAT, and cleaved caspase-3. In support, the histopathological examination of renal tissues was performed. UMB improves kidney function through regulation of renal serum biomarkers, with alleviations of histological abrasions induced by GNT. Besides, UMB downregulates renal protein expressions of ERK1/ERK2, TLR-4, and p38MAPK, with subsequent suppression of NF-kappa B-p65/NLRP-3 inflammasome and JAK1/STAT-3 pathways as well as cleaved caspase-3. In parallel, UMB induced IkB alpha upregulation. Collectively, UMB markedly amended all GNT-induced renal changes. These nephroprotective outcomes could be attributed to its ability to impede TLR-4/NF-kappa B-p65/NLRP-3 inflammasome and JAK1/STAT-3 pathways activation, as well as to its anti-inflammatory property.
COA
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Publication ()
Figure 1. Detection of G418 accumulation in cells using an indirect competitive gentamicin assay kit 
Figure 1. Structure of Gentamicin.
