Hormones & Inhibitors, etc.

Hormones & Inhibitors, etc. kdorfman Wed, 06/20/2018 - 21:25

By compound

By compound kdorfman Mon, 04/23/2018 - 17:49

ATP

ATP kdorfman Mon, 04/23/2018 - 17:50

10 mM ATP Stock
(from Maniotis)

  • 60 mg ATP in 8 mL H2O
  • pH to 7.0 with 0.1 M NaOH
  • Bring volume to 10 mL with H2O
  • Aliquot and freeze.

Angiotensin

Angiotensin kdorfman Thu, 04/26/2018 - 17:10

Sigma A9525 1mg/mL in water

Vasoconstrictor

Soluble in water 25 mg/mL (only use sterile water!)

MW: 1046.18 Asp-Arg-Val-Tyr-Ile-His-Pro-Phe

Subject to degradation in freezer storage - wear gloves!

Make 1 mg/mL stock (= 1.0462 mM)

Working concentration 1 µM

Dissolve 1:1000 in HBS

BP-A

BP-A kdorfman Mon, 03/22/2021 - 18:11

Bisphenol A

Sigma 239658

MW 228.29

100 mM stock in DMSO = 0.023g/mL

BP-S

BP-S kdorfman Mon, 03/22/2021 - 18:12

Sulfonyldiphenol

Sigma 103039

MW = 250.27

100 mM stock in DMSO = 0.025g/mL

Blebbistatin

Blebbistatin kdorfman Mon, 04/23/2018 - 17:49

Myosin inhibitor

(-)-Blebbistatin Sigma B05650-1mg

Info from Cayman:

A stock solution may be made by dissolving the (±)-blebbistatin in an organic solvent purged with an inert gas. (±)-Blebbistatin is soluble in organic solvents such as DMSO and dimethyl formamide (DMF). The solubility of (±)-blebbistatin in these solvents is approximately 10 mg/mL.

(±)-Blebbistatin is sparingly soluble in aqueous buffers. For maximum solubility in aqueous buffers, (±)-blebbistatin should first be dissolved in DMF and then diluted with the aqueous buffer of choice. (±)-Blebbistatin has a solubility of approximately 0.5 mg/ml in a 1:1 solution of DMF:PBS (pH 7.2) using this method. We do not recommend storing the aqueous solution for more than one day.

(±)-Blebbistatin is a selective cell-permeable inhibitor of non-muscle myosin II ATPases.1,2 It rapidly and reversibly inhibits Mg-ATPase activity and in vitro motility of non-muscle myosin IIA and IIB for several species (IC50 = 0.5-5.0 μM), while poorly inhibiting smooth muscle myosin (IC50 = 80 μM).3 Through these effects, blebbistatin blocks apoptosis-related bleb formation, directed cell migration and cytokinesis in vertebrate cells. Blebbistatin is inactivated by UV light,4 which may be particularly important in fluorescent cell imaging applications.

Note green fluorescent crystals seen in a 75µM solution.

Stock was 1 mg/mL in DMSO

44 µL 1mg/mL blebbistatin in DMSO in 1 mL non-CO2 medium

REDO

1 mg in 34 µL DMSO = ~100mM

MW = 292

Use at ~150 µM

Bradykinin

Bradykinin kdorfman Mon, 04/23/2018 - 18:56
  • 9-amino acid peptide chain: Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg

Bradykinin raises internal calcium levels

*Sigma B3259 - 1 mg septum bottle

  • MW = 1060.2

    1 mg/mL stock
    (= 1.0602 mM)

Add 1 mL H2O to the 1 mg in the bottle (NO CALCIUM! - some will be used for the calcium experiments in Lab 7.2)

Aliquots are 21.2 µL.

2 µM solution for use
(=2.12 µg/mL)

Brefeldin A

Brefeldin A kdorfman Tue, 11/12/2019 - 20:48

Fisher 00-4506 1000x solution

Keep in refrigerator

Description

Brefeldin A is an inhibitor of intracellular protein transport. Incubation of cells in culture with Brefeldin A leads to blockade of protein transport to the Golgi complex (GC) and accumulation of proteins in the endoplasmic reticulum (ER). Addition of Brefeldin A during the last hours of in vitro activation of cells results in enhanced detection of intracellular cytokines. Brefeldin A is effective for enhanced detection of a majority of mouse and human intracellular cytokines; however, it is advised that the investigators evaluate the use and efficacy of this reagent as well as other protein transport inhibitors such as Monensin in their specific assay system.

Applications Reported

The Brefeldin A Solution is supplied at 1000X working concentration in methanol. Prior to addition to the cell cultures, a 1:1000 dilution in the culture media should be made. The final working concentration of the Brefeldin A (at 1x) is 3.0 ug/ml.

Caffeine

Caffeine kdorfman Tue, 04/13/2021 - 15:25

MW 194.19

Soluble 10 mg/mL in water (with heat and stirring)

Make a 50mM (=0.05M) solution: 9.71mg/mL

Cannabinol

Cannabinol kdorfman Fri, 05/11/2018 - 16:11

Tetrahydrocannabinol solution 1 mg/mL in methanol

Sigma T464

store at 4C

Concanavalin A

Concanavalin A kdorfman Mon, 04/23/2018 - 18:21
  • Sigma C5275 5 mg
  • Plant mitogen
  • Lectin (carbohydrate-binding protein) extracted from the jack-bean, Canavalia ensiformis
  • Con A binds specifically to α-Mannose, α-Galactose structures found in sugars, glycoproteins and glycolipids

  • Make stock 5 mg/mL solution (add 1 mL sterile PBS to bottle – gently rotate to mix)

  • Keep sterile - work in sterile hood

  • aliquot 100 µL into microfuge tubes  (cannot take repeated freeze-thaw cycles)

Con A coverslips

Con A coverslips kdorfman Sun, 05/24/2020 - 15:38

*Requirements:**      * Acid washed coverslips stored in 100% ethanol.

  • ConA 0.5 mg/mL. 

  • Large petridishes to store the coverslips.    

  Procedure:

  • Flame the acid-washed coverslips and cool for 10 sec.    (Make 9-­‐14 coverslips     at a    time    to save time.)

  • Place the coverslips on top of the    large petri dish.     

  • Thaw the conA from the freezer and dilute to 0.5 mg/mL 
  • apply around 400uL the first two coverslips 

  • Wait for 10 sec. 

  • Tilt the coverslips and suck out the excess ConA 

  • Repeat the same for all the coverslips. 

  • Cover the petri dish with another lid of petri dish and allow a small opening to dry.

  • If possible,use a vacuum pump  to dry it faster.   

The process usually takes 15 minutes and drying takes half an hour. 

Storage:

  * In a petri dish, place the coverslips on top of a clean kimwipe with the conA surface facing up.

  • Cover with the lid. 10 min before needed, UV sterilize them.

  • Alternatively, UV sterilize all of them after drying and open the dishes only in aseptic conditions or inside hood.       

Con A dishes

Con A dishes kdorfman Sun, 05/24/2020 - 15:37

Concanavalin-coated coverslip dishes

  • Start with 5 mg/mL ConA stock solution

  • Keep sterile - work in sterile hood

  • aliquot 100 µL into microfuge tubes  (cannot take repeated freeze-thaw cycles)

  • Coat coverslip bottom dishes with either:

    • 0.75 mL 0.1 mg/mL:  0.015 mL (5 mg/mL) + 0.735 mL PBS
    • 0.75 mL 0.5 mg/mL:  0.075 mL (5 mg/mL) + 0.675 mL PBS
  • Incubate the coverslip dishes for 1 hour 37˚C 

  • Remove the solution

  • rinse in PBS 

  • Air dry the dish.

Cyclin Inhibitor

Cyclin Inhibitor kdorfman Wed, 04/25/2018 - 19:55

CKD1 inhibitor IV RO3306

1 mM in DMSO

A cyclin-dependent kinase inhibitor protein inhibits cyclin-dependent kinase. Several function as tumor suppressor proteins. Cell cycle progression is negatively controlled by cyclin-dependent kinases inhibitors (called CDIs, CKIs or CDKIs). CDIs are involved in cell cycle arrest at the G1 phase.

Cyclohexiamide

Cyclohexiamide bcrcstaff Thu, 03/21/2019 - 13:28

Protein synthesis inhibitor

Fisher AC357420010

MW =  281.35 g/mol

C15H23NO4

powder

Cytochalasin D

Cytochalasin D kdorfman Mon, 04/23/2018 - 18:21

MW = 507.62
Sigma C2618: 200 µL 5 mg/mL (~10 mM) in DMSO
Sigma C8273: 5 mg. Dissolve in 1 mL DMSO

  • alkaloid mycotoxin produced by Helminthosporium and other molds.
  • disrupts actin microfilaments
  • binds to F-actin polymer
  • prevents polymerization of actin monomers
  • Aliquot 10 µL 10 mM
  • Add 90 uL DMSO (makes 1 mM)
  • Dilute this 1:1000 into medium (makes 1uM)
  • Dilute the 1 uM to desired working concentration

  • Store in the dark, in the freezer

  • Adding 490 µL of non-CO2 medium makes 500 µL of 200 µM

  • Final dilution should contain no more than 0.1% DMSO

  • 250 nM in the dish should give an effect

  • 15 - 30 min

Fibronectin

Fibronectin kdorfman Thu, 04/26/2018 - 17:15

Sigma F1141

Attachment factor

GSA 10

GSA 10 kdorfman Thu, 01/28/2021 - 18:39

Smo (Smoothened) receptor* agonist

Tocris 4918 10 mg

MW = 459.94

Make a 5 mM stock solution in 4.34 mL DMSO (heat to 50C to dissolve)

*Non-classical G-protein-coupled receptors that belong to the Frizzled family. Smoothened receptors lack the ability to directly interact with their endogenous ligand, Hedgehog (Hh). In the resting state, Smo receptors are bound to patched (Ptc).

Does not recognize the classic cyclopamine (Cat. No. 1623) binding site. Does not promote Smo translocation to the primary cilium; is strongly potentiated by forskolin and cholera toxin. Displays anti-adipogenic effects in vitro, mediated by a non-canonical Hedgehog signaling pathway. Promotes differentiation of multipotent mesenchymal progenitor cells into osteoblasts.

H-89

H-89 kdorfman Mon, 03/22/2021 - 18:08

H 89 hydrochloride Tocris 2910

1 mg

MW 532.79

1 mg/mL in DMSO

HU (Hydroxyurea)

HU (Hydroxyurea) kdorfman Wed, 01/06/2021 - 18:29
ingredient working conc MW stock
HU 0.2 M 76 Koshand lab 2M in H2O

Hydroxyurea: (Sigma H8267-1g)

  • To make 50 mL YPD with 0.2M HU
  • vi = (0.2M * 50 mL)/2M = 5 mL

  • 100µL 2M stock per mL medium

  • Buy 1 g, make 6.6 mL 2M stock

  • MW = 76, so 1 g = 1/76 mol = 0.0132 mol
  • vol = (1000 mL/2 mol) * 0.0132 mol = 6.6 mL

See yeast recipes for QBoC https://wahoo.cns.umass.edu/node/554/

Hydrocorisone

Hydrocorisone kdorfman Mon, 03/22/2021 - 18:06

Fisher A16292 (Alfa Aesar) 1 g

1 mg/mL in 100% ethanol.

Importazole

Importazole kdorfman Thu, 05/10/2018 - 14:25

401105-10mg Calbiochem

Importazole is an inhibitor of importin-β transport receptors.

During interphase, the transport receptor importin-β carries cargoes into the nucleus, where RanGTP releases them. Importazole somehow disrupts the importin/RAN interaction. Importazole is selective among other transporters. Compounds are imported into, but not out of, cells.

A cell-permeable diaminoquinazoline compound that selectively blocks importin-β-mediated nuclear import of NLS bearing cargos in a reversible manner.

Soluble in DMSO (25 mg/ml; clear, colorless solution)

5 mg/mL ailquots in freezer

Ionomycin

Ionomycin kdorfman Mon, 04/23/2018 - 18:47

Raises intracellular calcium levels

Fisher or Invitrogen (Life Technologies) I24222

1 mM in DMSO or EtOH. 1 mg makes 1.34 mL stock.

3 µM Ionomycin in HBS
Dilute stock 1:333 in HBS: 3 µL/1 mL

24 µL in 8 mL, aliquot 800 µL

They only need 750µL, because they do one experiment in HBS, then two in Ca-free HBS.

3 µM Ionomycin in Ca-free HBS
Dilute stock 1:333 in Ca-free HBS: 3 µL/1 mL
45 µL in 15 mL

Takes a long time to thaw. Vortex.

Aliquot 1.55 mL so they can do two 750 µL experiments

Jasplakinolide

Jasplakinolide kdorfman Thu, 04/26/2018 - 17:44

(out of stock 2018)

  • Jasplakinolide – Santa Cruz biochemical, product sc202191 F
    • inducer of actin polymerization & stabilization; inhibits actin filament disassembly
    • F-actin probe
    • MW = ~710
    • powder is stable frozen over a year
    • stock is stable 3-4 mo at -20C
    • Make 1 mM stock with 50 µg in 70 µL DMSO
    • Add 5 mL non-CO2 medium to make ~14 µM
    • 50 nM to 5 μM working range
    • minutes to hours incubation time

LDL

LDL kdorfman Fri, 05/11/2018 - 16:29

Low-density lipoprotein from human plasma, Dil complex (Dil LDL)

1 mg/mL

200 uL

DO NOT FREEZE

Life Technologies L3482

Laminin

Laminin kdorfman Thu, 04/26/2018 - 18:16

Laminin from mouse BD354232

1 mL 1 mM

Cell adhesion

Latrunculin

Latrunculin kdorfman Thu, 04/26/2018 - 17:46

Alexis Biochemicals 350-036-C100 Catalog # t110

Inhibits actin polymerization and disrupts microfilament organization, as well as microfilament-mediated processes

Reported to be 10 to 100-fold more potent than the cytochalasins. May act more slowly, though. Lots of variation between cell lines.

Inactivated by FBS. (Rinse medium off before treatment.)

MW = 395.5

Soluble in DMSO and ethanol.

Store in freezer.

Active concentration range: 90 nm ( =~0.1µM) to 2.5 µM

(2.5 µM = 25 x 100 nm)

Stock is 1mm

µL 1mM stock µL serum-free buffer or medium µM final concentration
1 * 99 10
1 399 2.5
1 999 1

*Use this to make further dilutions

ML-7

ML-7 kdorfman Wed, 04/25/2018 - 19:42

ML-7 Sigma, 12764.

selective myosin light chain kinase inhibitor

  • soluble 10mg/mL in 50% EtOH
  • MW = 452.74
  • 5 mg in bottle
  • add 1100 µL to make 10 mM solution
  • used at ~50 µM (1 µL/1mL) (Pat says 75 µM)
  • 75 µL 10 mM + 5 mL non-CO2 medium makes 150 µM
  • make 15 µL aliquots of 10 mM. Add 985 µL medium to make 150 µM

Melanocyte Stimulating Hormone

Melanocyte Stimulating Hormone kdorfman Wed, 04/25/2018 - 20:48

Sigma M4135 1 mg

Molecular Weight 1664.88

From Sigma: Amino Acid Sequence:

Ac-Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-Pro-Val-NH2

Pituitary hormone which causes darkening skin pigmentation from amphibians to humans. In mammals, it can also have behavioral effects on learning, attention, and memory.

Hormone that stimulates melanogenesis; facilitates learning and memory; affects inflammatory and immune responses and peripheral nerve regeneration.

α-Melanocyte-stimulating hormone (α-MSH) acts as an anti-inflammatory agent via down regulating the production and activity of the pro-inflammatory cytokines interleukin-1 (IL-1), tumor necrosis factor (TNF)-α and IL-6 expressed in various cells of the immune system. It also controls the nitric oxide production associated with inflammation. α−MSH inhibits nuclear factor-κB (NF-κB)-dependent gene transcription and NF-κB pathway induced by TNF and other inflammatory agents. This activity of α-MSH is mediated through the production of cyclic adenosine monophosphate (cAMP) and activation of protein kinase A (PKA) enzyme. α–MSH functions as a potent therapeutics for various conditions resulted through NF-κB activation including, inflammatory diseases, human immunodeficiency virus (HIV) replication in AIDS (acquired immunodeficiency syndrome), and septic shock.[2] α-MSH has an essential role to play in melanin production in animals. α-MSH regulates development of several skin diseases, including cutaneous inflammation and hyper-proliferative skin diseases.[3] Linkage

Derived from ACTH 1-13 Other Notes

Lyophilized from 0.1% TFA in H2O Packaging

1, 5 mg in glass bottle Application

α-Melanocyte stimulating hormone (α–MSH) has been used for following studies: • it has been Intracerebroventricularly (icv) injected to mice for behavioral studies.[1] • to determine the effect of α−MSH on growth of stationary Nb 2 node lymphoma cell cultures.[4] • to determine the effect of α−MSH on leptin secretion in the primary cultures of differentiated adipocytes.[5] • α−MSH promoted melanin production in the B16-F1 cells from murine melanoma cell line.[6] General description

α-Melanocyte-stimulating hormone (α-MSH) is a tridecapeptide, mostly produced by the cells in the brain, pituitary and circulation.[2] Pro-inflammatory cytokines or UV light induced epidermal cells such as keratinocytes and melanocytes synthesize and discharge α–MSH. Poopiomelanocortin (POMC) acts as a precursor for α-Melanocyte-stimulating hormone (α-MSH) production.[3]

Monastrol

Monastrol kdorfman Thu, 05/10/2018 - 15:17

Inhibits kinesin 5

200 mM in DMSO

Final concentration 200 uM

5.5 uL aliquots

Monensin

Monensin kdorfman Wed, 04/25/2018 - 21:25

Monensin sodium salt

Sigma M5273

soluble in methanol 50 mg/mL

insoluble in water

soluble in DMSO, ethanol

ionophore which disrupts the structure of the Golgi apparatus and inhibits vesicular transport in eukaryotic cells

inhibits transition from G1 to S

antiprotozoal, antibacterial, or antifungal agent

MW 692.9

1 µM = half-maximal inhibitory effect

Nerve Growth Factor

Nerve Growth Factor kdorfman Fri, 05/11/2018 - 16:16

Nerve Growth Factor 2.5S

Harlan.com Cat no. 5025

From Sigma: 2.5S subunit of nerve growth factor-7S (NGF-7S) is essentially the β-subunit when isolated from male mouse submaxillary glands under initially dissociative conditions by a modification of the method of Bocchini and Angeletti.

Stored at 4C

Nocodazole

Nocodazole kdorfman Mon, 04/23/2018 - 17:51

Prevents polymerization of microtubules. Prevents cells from entering metaphase.

*Acros 358240100, 10 mg

*Soluble in DMSO

*Dilute stock in medium to treat cells.

*Make a 10 mg/mL (=33mM) stock: Add 1 mL DMSO to 10 mg in bottle.

  • Aliquot 3 µL 33 mM per tube.

*Working concentration range is usually 100nM - 30 µM.

  • add 97 µL DMSO to tube to make 100 µL of 1 mM

  • 33 µL 1mM + 10 mL medium = 3.3 µM

Plant hormones

Plant hormones bcrcstaff Thu, 03/21/2019 - 14:49
Compound function Form Room storage stock Cat #
ABA Plant hormone involved in growth and stress response powder 362A fridge 50 mM in EtOH Fisher AC133480010
ABA powder soluble 20 mg/mL in DMSO, ethanol 362A freezer PlantMedia 30631017-1
soluble in methanol
Ascorbic acid Enzyme cofactor Powder 362A RT Fisher S26184
Caffeine Growth inhibitor Powder 362A RT Fisher ICN15011483
Chitosan medium viscosity Component of fungal cell walls and arthropod exoskeletons Powder 362A RT Bio-World 40300161-1
Cycloheximide Protein synthesis inhibitor Powder 362A RT Fisher AC357420010
Gibberellic acid breaks seed dormancy 13 mg/mL (=37.5mM) solution 362A 4C PlantMedia 30631025-1
IAA auxin growth regulator
stimulates root growth
solid 362A 4C 100 mM in EtOH PlantMedia 30631010-
Imidacloprid (pestanal) Systemic insecticide neat 362A dessicator Sigma 37894
kinetin Plant cytokinin Powder 362A RT Sigma K0753
Lactic acid Plant growth stimulator 69% soln aq 362A dessicator Fisher AC250300100
Methyl jasmonate Pathogen response Liq 362A RT PlantMedia30631015-3
Nicotine Plant defense chemical liq 362A dessicator Fisher AC181420050
Salicylic acid Response to abiotic stress Powder 362A RT 100 mM in EtOH Fisher S25515
Selenium varies Powder 362A RT Strem 93-3416

Poly-L-Lysine

Poly-L-Lysine kdorfman Thu, 04/26/2018 - 17:04

Non-specific attachment factor for cells; use it to coat coverslips.

Sigma P1524

  1. Add 50 ml of sterile tissue culture grade water to 5 mg of poly-lysine.
  2. Aseptically coat culture surface with 1 ml per 25 cm2 of solution. Rock gently to ensure even coating of the culture surface.
  3. After 5 minutes, remove solution by aspiration and thoroughly rinse surface with sterile tissue culture grade water.
  4. Allow to dry at least two hours before introducing cells and medium. If glassware or slides must be sterilized after coating with poly-lysine, γ-irradiation is recommended instead of autoclaving.

Can also buy poly-lysine coated slides: Fisher 6776215

Purmorphamine

Purmorphamine kdorfman Thu, 01/28/2021 - 18:25

Tocris 4551 10 mg

MW = 525.12

Make a 10 mM stock in DMSO in 1.9 mL DMSO

STLC

STLC kdorfman Mon, 04/23/2018 - 17:51
  • STLC Sigma 164739 (Fisher 50-703-1833), MW = 363.47

(+)-S-Trityl-L-cysteine is a cell-permeable selective inhibitor of mitotic kinesin Eg5 and ATPase activities.

KIF11 (also known as kinesin-5 and Eg5) is a homotetramer which cross-links anti-parallel microtubules in the mitotic spindle to maintain spindle bipolarity. The motor domain or motor head is at the N-terminus and performs ATP hydrolysis and binds to microtubules. (https://en.wikipedia.org/wiki/Kinesin_family_member_11)

  • Make 100 mM stock:

    • 1 g in bottle.
    • Mix with 27.5 mL DMSO
    • heat to 65C (~10 min), vortex
    • if necessary, filter sterilize to remove insoluble particles.
    • save as 1 mL aliquots in freezer. (dilute 1:10,000 to use) (dilute 1:100 to make 1 mM)
  • Make 1 mM stock from the 100 mM stock

    • 10 µL 1mM stock
    • 990 µL DMSO
    • vortex
    • it will crystallize in the refrigerator. Warm and vortex to redissolve it.
  • Make 10 µL aliquots of 1 mM stock. Freeze

    • label says to add 990 µL medium to make 10 µM working solution

Taxol

Taxol kdorfman Mon, 04/23/2018 - 17:51

Taxol (20 µM final)

Fisher 109710 Paclitaxel.
Sigma T7402 1mg
MW = 853.91

  • Stabilizes the microtubule polymer
  • Protects microtulues from disassembly.
  • Prevents metaphase spindle configuration
  • Blocks progression of mitosis and prolonged activation of the mitotic checkpoint triggers apoptosis or reversion to the G-phase of the cell cycle without cell division.

  • soluble in DMSO, not water.

  • Add 0.117 mL DMSO to make a 0.01M (=10 mM) stock solution from the 1 mg powder

  • Aliquot 10 µL 10 mM (if it precipitates, add 10 uL DMSO and vortex, then add 480 uL medium or buffer)

    • add 490 µL to make 200µM
    • dilute into medium from there

Thyroid hormones

Thyroid hormones kdorfman Fri, 09/06/2019 - 18:38

for Fish

5 mL per well for treatment

PTU (inhibitor) Sigma p3755
MW 170.2
Solubility:

  • 50 mg/mL in 1M NaOH
  • 16 mg/mL in alcohol
  • 1 mg/mL in RT water
  • 10 mg/mL in boiling water

In order to get the following doses, best to make it in hot fish water; otherwise, the NaOH or EtOH will be too concentrated in the final dilution.

  • dose 1: 0.5 mM

  • dose 2: 1.0mM

MW of PTU:
Concentration: M
to make: mL
add: g PTU

L-T4 (thyroxin) Sigma T2376
MW: 776.87
solubility:

  • 50 mg/ml in 4M NH4OH in MeOH
  • 0.5 mg/mL in 1:5 DMSO:PBS

Solution is good for ~ 1 day.

Make a 1 mM stock in 1M NaOH. Takes a long time. Vortex.

Priya's recipe: 1 mg/mL in 1M NaOH (~25.7 uM stock)

Working concentrations

  • dose 1: 100 nM (=0.1 uM)

  • dose 2: 300 nM (=0.3 uM)

MW of T4:
Concentration: M
to make: mL
add: g T4

Trichostatin A

Trichostatin A kdorfman Wed, 04/25/2018 - 19:52

Cayman 89730

10 mM

Histone deacetylase Inhibitor

Trichostatin A (TSA), an antifungal antibiotic produced by Streptomyces hygroscopicus, is a potent and specific inhibitor of histone deacetylases (HDACs), which are overexpressed in various cancers and closely correlate with oncogenic factors.

Trichostatin A is active at nanomolar concentrations in mammalian cells. By suppressing the activity of HDACs, it leads to increased histone acetylation, thereby causing highly acetylated histones to accumulate in the cell [2]. This in turn induces enhanced expression of specific genes that elicit extensive cellular morphologic and metabolic changes such as growth arrest, differentiation and apoptosis. At submicromolar concentrations Trichostatin A has been shown to induce apoptosis in diverse cancer cells while exhibiting very low toxicity to normal cells.

TSA is sparingly soluble in aqueous buffers. For maximum solubility in aqueous buffers, TSA should be directly dissolved in 0.1 M HCl (1.3 mg/ml) and then neutralized with PBS (pH 7.2) to achieve the desired concentration or pH. Approximately 0.7 ml of PBS (pH 7.2) is required to neutralize 1 ml of the acidic solution. We do not recommend storing the aqueous solution for more than one day.

Valinomycin

Valinomycin kdorfman Wed, 04/25/2018 - 21:03

Potassium Ionophore, MW: 1111.32

Sigma V0627

Insoluble in water

stock is 10 mg/mL in DMSO

  • cyclododecadepsi-peptide ionophore antibiotic.
  • potassium ionophore that transports K+ across biological and artificial lipid membranes.
  • can induce K conductivity in cell membranes
  • uncouples oxidative phosphorylation,
  • induces apoptosis in murine thymocytes, and in pre-B cell.
  • inhibits NGF-induced neuronal differentiation
  • antagonizes ET-induced vasoconstriction.
  • Useful in studies of K transport in mitochondria.

Vasopressin

Vasopressin kdorfman Mon, 04/23/2018 - 17:50

(=Anti-diuretic hormone)

causes calcium oscillations

1 mM stock in water
(NO CALCIUM, so stock can be used for Ca-free solutions in Lab 7.2)

1 mg in bottle, MW = 1084
Add 0.92 mL to make 1mM stock

1 µL aliquots in freezer


1 µM dilution to make student solutions

1 µL 1 mM in 999 µL water

(NO CALCIUM, so stock can be used for Ca-free solutions in Lab 7.2)