200 µg, unless otherwise indicated in figure legends, of polyclonal rat IgG (BioXcell; isotype) or anti-LAG-3 (clone C9B7W, BioXcell) and/or anti-PD-L1 (10F.9G2, BioXcell) or anti-PD-1 (RMP1-14, BioXcell) blocking Abs were injected i.v. into blood-stage parasitized mice beginning on day 9 or 12 after infection (as indicated) and every 3 days until day 15 or 18 as indicated. Antibodies which contained endotoxin levels above 3 EU/ml (Kinetic-QCL Kinetic Chromogenic LAL Assay, Lonza) were depleted of endotoxin with the ToxinEraser Endotoxin Removal kit (GenScript) following manufacturer’s instructions.
Kinetic qcl kinetic chromogenic lal assay
Manufactured by Lonza
Sourced in United States
The Kinetic-QCL Kinetic Chromogenic LAL Assay is a laboratory equipment product designed to detect and quantify endotoxin levels in various samples. It utilizes a kinetic chromogenic method to measure the endotoxin concentration. The assay provides a quantitative analysis of endotoxin levels in the tested samples.
Automatically generated - may contain errors
Lab products found in correlation
Toxineraser endotoxin removal kit, by GenScript (1 mentions)
Anti lag 3 clone c9b7w, by BioXCell (1 mentions)
Rmp1 14, by BioXCell (1 mentions)
Hiload 26 600 superdex 200 pg column, by GE Healthcare (1 mentions)
Capto q, by GE Healthcare (1 mentions)
Capto, by Cytiva (1 mentions)
Mycoalert mycoplasma detection kit, by Lonza (1 mentions)
C57bl 6, by Charles River Laboratories (1 mentions)
Scid mice, by Charles River Laboratories (1 mentions)
5 protocols using kinetic qcl kinetic chromogenic lal assay
1
Malaria Immunotherapy Protocol
2
Purification of Fusion Protein TFF2-CTP-Flag
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The supernatant was harvested by centrifugation at 500 × g for 15 min at 4 °C. The soup was passed through 0.45 μM filter to remove any remaining cells debris. Then the soup was concentrated to 1/10th of starting volume using 5000 Da cutoff vivaflow 50 cassettes (Sartorius). Then the concentrated soup was diluted to ten times using Equilibration buffer (20 mM Tris-HCl, 20 mM NaCl pH 8.0). Fusion TFF2-CTP-Flag protein from this diluted soup was purified using anion-exchange chromatography (Capto Q, GE life sciences). The bound protein was eluted using a salt gradient through elution buffer (20 mM Tris-HCl, 1 M NaCl pH 8.0). The eluate was then concentrated, filtered and applied to a size exclusion chromatography (HiLoad 26/600 Superdex 200 pg column, GE Healthcare, England) equilibrated with DPBS buffer. The fusion TFF2-CTP-Flag protein containing fractions were pooled, concentrated, filtered and checked for endotoxin levels (Kinetic-QCL™ Kinetic Chromogenic LAL Assay, Lonza).
3
Serum Endotoxin Levels in Extreme Feed Efficiency
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On the last day of the feeding trial (day 70), blood was collected from the jugular vein of all animals in vacuum tubes containing clot activator. The tubes were centrifuged (4 °C, 3500 rpm) for 15 minutes to separate the serum. Serum aliquots were stored in −80 °C freezer until analysis. Serum endotoxin dosage of 10 extreme animals (more animals than the proteome analysis but still high or low extreme animals for FE) of each FE group (HFE and LFE) was performed by using the Kinetic-QCL Kinetic Chromogenic LAL Assay (Lonza, 50-650U). For this purpose, the samples were preheated at 70 °C for 15 minutes and used pure or diluted 1:1 or 1:4 in order to dilute the endotoxins to reach the detection range of the kit for absolute quantification. One sample presented no absolute quantification, being indicated as <0.01 EU/ml and the value of 0.01 EU/ml was assigned to it for the statistical test. In addition, two samples presented values ≥2.5 SD from the mean of all samples and were withdraw from the analysis. High and low FE groups were tested by Student’s t-test.
4
Cell Contamination Screening Protocols
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The cells were tested for mycoplasma contamination using a MycoAlert Mycoplasma Detection Kit (LT07-218; Lonza; USA) and for endotoxin contamination using a Kinetic-QCL™ Kinetic Chromogenic LAL Assay (50-650NV; Lonza; USA) according to the manufacturers' instructions.
5
Antibody Characterization for In Vivo Studies
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CD-1, C57BL/6, and SCID mice used in Toronto were from Charles River Laboratories. Monoclonal anti-platelet antibody (MWReg30, rat IgG1) was purchased from BD Biosciences. The 30-F1 (rat IgG2c) antibody recognizing mouse RBC was from BioLegend. Antibodies 34-1-2s (rat IgG2a) and Ter119 (rat IgG2b) were from Bio X Cell. Mouse monoclonal anti-type II collagen 5 clone antibody mixture was purchased from Chondrex. The control rat IgG used in Fig. 1 (ChromPure) was from Jackson ImmunoResearch. The Ter119, deglycosylated Ter119, mouse IgG1 Ter119, and mouse IgG2a Ter119 used in Figs. 1A, 2 (E andF), and 3 to 8 and figs. S1 to S3 were all still available at the time of writing the manuscript and were tested together in a single batch for endotoxin levels using the Lonza "Kinetic-QCL" Kinetic Chromogenic LAL Assay (catalog no. 50-650 U). These antibodies either had an endotoxin level below the limit of detection of the assay or were present at a level below the United States Pharmacopeia (USP)-recommended endotoxin limit of 5 EU/kg body weight for parenteral drugs, as well as below the same recommended limit for preclinical research (72, 73) . The Ter119 used in Figs. 1A and7 (the temperature experiments) had an endotoxin level below the limit of detection providing a maximal theoretical in vivo dose of <0.76 EU/kg body weight.
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