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Bravo instrument

Manufactured by Agilent Technologies
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The Bravo instrument is a versatile liquid handling platform designed for automated sample preparation and processing in a laboratory setting. It features precise and accurate liquid handling capabilities to support a wide range of applications.

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Lab products found in correlation

Rneasy micro kit, by Qiagen (4 mentions) Sybr green master mix, by Roche (3 mentions) Maxima reverse transcriptase, by Thermo Fisher Scientific (2 mentions) Palindromic forked adapters, by Integrated DNA Technologies (2 mentions) Palindromic forked adapters, by Roche (2 mentions) Lightcycler 480 sybr green 1 master mix, by Roche (1 mentions) Maxima first strand cdna synthesis kit for rt qpcr, by Thermo Fisher Scientific (1 mentions) Lightcycler 480 2, by Roche (1 mentions) Maxima first strand cdna synthesis kit, by Thermo Fisher Scientific (1 mentions) 96 reaction kits, by Roche (1 mentions) Paired end adapter, by Illumina (1 mentions) Paired end adapters, by Integrated DNA Technologies (1 mentions)

Spelling variants of this product

Bravo pipetting robot instrument (3 citations)

6 protocols using bravo instrument

1

Quantitative RT-PCR for Gene Expression

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RNA was extracted from cells with the RNeasy Micro kit (QIAGEN). Reverse transcription was performed with the Maxima First Strand cDNA Synthesis Kit for RT-qPCR (Thermo Fisher Scientific). cDNA was prepared with LightCycler 480 SYBR Green I Master mix (Roche) using the Bravo instrument (Agilent) and analyzed on a LightCycler 480 instrument using a 2-step protocol with a 60°C annealing/elongation step. All samples were run in technical triplicates, and the average Ct-values were used for calculations. Data are represented using the DDCt method. All fold changes are calculated as the average fold change based on 2 different housekeeping genes (ACTB and GAPDH). The primers used are described in Table S1 (Integrated DNA Technologies).
Adler A.F., Cardoso T., Nolbrant S., Mattsson B., Hoban D.B., Jarl U., Wahlestedt J.N., Grealish S., Björklund A, & Parmar M. (2019). hESC-Derived Dopaminergic Transplants Integrate into Basal Ganglia Circuitry in a Preclinical Model of Parkinson’s Disease. Cell Reports, 28(13), 3462-3473.e5.
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2

RNA Isolation and Quantitative RT-PCR

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Total RNAs were isolated on day of analysis using the RNeasy Micro Kit (QIAGEN#74004) according to manufacturer instructions. Approximately 1 μg of RNA was reverse transcribed using Maxima First Strand cDNA Synthesis Kit (Thermo Fisher 2#K1642Invitrogen). cDNA was prepared together with SYBR Green Master mix (Roche#04887352001) using the Bravo instrument (Agilent) and analyzed by quantitative PCR on a LightCycler 480 II instrument (Roche) using a 2-step protocol with a 95 °C, 0.5 min denaturation step followed by a 60 °C, 1 min annealing/elongation step for 40 cycles in total. All quantitative RT-PCR (qRT-PCR) samples were run in technical triplicates, analyzed with the ΔΔCt-method, normalized against the two housekeeping genes ACTB (actin-beta) and GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and results are given as a fold change of expression over undifferentiated hPSCs. Details and list of primers are reported in Supplementary Table S1.
Nilsson F., Storm P., Sozzi E., Hidalgo Gil D., Birtele M., Sharma Y., Parmar M, & Fiorenzano A. (2021). Single-Cell Profiling of Coding and Noncoding Genes in Human Dopamine Neuron Differentiation. Cells, 10(1), 137.
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3

RNA Isolation and qRT-PCR Analysis

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Total RNAs were isolated using the RNeasy Micro Kit (QIAGEN, #74004) and reverse transcribed using random hexamer primers and Maxima Reverse Transcriptase (Thermo Fisher, #K1642Invitrogen). cDNA was prepared with SYBR Green Master Mix (Roche, #04887352001) using the Bravo instrument (Agilent) and analyzed by quantitative PCR on a LightCycler 480 supplier using a 2-step protocol with a 60 °C annealing/elongation step. All quantitative reverse transcriptase PCR (qRT-PCR) samples were run in technical triplicates and results are given as fold change over undifferentiated hPSCs. Details and list of primers are reported in Supplementary Table 1.
Fiorenzano A., Birtele M., Wahlestedt J.N, & Parmar M. (2021). Evaluation of TH-Cre knock-in cell lines for detection and specific targeting of stem cell-derived dopaminergic neurons. Heliyon, 7(1), e06006.
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4

Optimized DNA Library Preparation for Sequencing

Cited 1 time
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DNA libraries for massively parallel sequencing were generated as previously described32 (link) with the following modifications: the initial genomic DNA input into the shearing step was reduced from 3 μg to 10–100 ng in 50 μl of solution. For adaptor ligation, Illumina paired-end adapters were replaced with palindromic forked adapters (purchased from Integrated DNA Technologies) with unique 8-base index molecular barcode sequences included in the adaptor sequence to facilitate downstream pooling. With the exception of the palindromic forked adapters, all reagents used for end repair, A-base addition, adaptor ligation, and library enrichment PCR were purchased from KAPA Biosciences in 96-reaction kits. In addition, during the post-enrichment solid phase reversible immobilization bead cleanup, elution volume was reduced to 20 μl to maximize library concentration, and a vortexing step was added to maximize the amount of template eluted from the beads. Libraries with concentrations above 40 ng μl−1, as measured by a PicoGreen assay automated on an Agilent Bravo instrument, were considered acceptable for hybrid selection and sequencing.
Taylor-Weiner A., Zack T., O’Donnell E., Guerriero J.L., Bernard B., Reddy A., Han G.C., AlDubayan S., Amin-Mansour A., Schumacher S.E., Litchfield K., Turnbull C., Gabriel S., Beroukhim R., Getz G., Carter S.L., Hirsch M.S., Letai A., Sweeney C, & Van Allen E.M. (2016). Genomic evolution and chemoresistance in germ-cell tumours. Nature, 540(7631), 114-118.
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5

Quantitative RT-PCR Analysis of hPSCs

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Total RNAs were isolated using the RNeasy Micro Kit (QIAGEN#74004) and reverse transcribed using random hexamer primers and Maxima Reverse Transcriptase (Thermo Fisher #K1642, Invitrogen). cDNA was prepared together with SYBR Green Master mix (Roche#04887352001) using the Bravo instrument (Agilent) and analyzed by quantitative PCR on a LightCycler 480 device using a 2-step protocol with a 60 °C annealing/elongation step. All quantitative RT-PCR (qRT-PCR) samples were run in technical triplicates and the results are given as fold change over undifferentiated hPSCs using each of the two housekeeping genes for normalization (ACTB and GAPDH). Details and list of primers are reported in Supplementary Table 1.
Fiorenzano A., Sozzi E., Birtele M., Kajtez J., Giacomoni J., Nilsson F., Bruzelius A., Sharma Y., Zhang Y., Mattsson B., Emnéus J., Ottosson D.R., Storm P, & Parmar M. (2021). Single-cell transcriptomics captures features of human midbrain development and dopamine neuron diversity in brain organoids. Nature Communications, 12, 7302.
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6

Optimized DNA Library Preparation for Massively Parallel Sequencing

Cited 1 time
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DNA libraries for massively parallel sequencing were generated as previously described (43 (link)) with the following modifications: the initial genomic DNA input into the shearing step was reduced from 3 μg to 10-100 ng in 50 μL of solution. For adapter ligation, Illumina paired-end adapters were replaced with palindromic forked adapters (Integrated DNA Technologies) with unique 8-base index molecular barcode sequences included in the adapter sequence to facilitate downstream pooling. With the exception of the palindromic forked adapters, all reagents used for end repair, A-base addition, adapter ligation, and library enrichment PCR were purchased from KAPA Biosciences in 96-reaction kits. In addition, during the post-enrichment solid phase reversible immobilization (SPRI) bead cleanup, elution volume was reduced to 20 μL to maximize library concentration, and a vortexing step was added to maximize the amount of template eluted from the beads. Libraries with concentrations above 40 ng/μL, as measured by a PicoGreen assay automated on an Agilent Bravo instrument, were considered acceptable for hybrid selection and sequencing.
Giltnane J.M., Hutchinson K.E., Stricker T.P., Formisano L., Young C.D., Estrada M.V., Nixon M.J., Du L., Sanchez V., Ericsson P.G., Kuba M.G., Sanders M.E., Mu X.J., Van Allen E.M., Wagle N., Mayer I., Abramson V., Gómez H., Rizzo M., Toy W., Chandarlapaty S., Mayer E.L., Christiansen J., Murphy D., Fitzgerald K., Wang K., Ross J.S., Miller V.A., Stephens P.J., Yelensky R., Garraway L., Shyr Y., Meszoely I., Balko J.M, & Arteaga C.L. (2017). Genomic profiling of ER+ breast cancers after short-term estrogen suppression reveals alterations associated with endocrine resistance. Science translational medicine, 9(402), eaai7993.
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